In the course of infection, the detection of pathogen-associated molecular patterns by specialized pattern recognition receptors in the host leads to activation of the innate immune system. Whereas the subsequent induction of adaptive immune responses in secondary lymphoid organs is well described, little is known about the effects of pathogen-associated ...

Form I Rubisco (ribulose 1,5-bisphosphate carboxylase/oxygenase), a complex of eight large (RbcL) and eight small (RbcS) subunits, catalyses the fixation of atmosphericCO2 in hotosynthesis. The limited catalytic efficiency of Rubisco has sparked extensive efforts to re-engineer the enzyme with the goal of enhancing agricultural productivity. To facilitate ...

Protein function such as catalytic activity or molecular recognition is tightly coupled to the conformation and dynamics. Since protein conformations may be controlled by forces, diverse active and passive mechanisms have evolved that allow biological systems to respond to mechanical signals. However, forces act in a predetermined direction on these ...

The p53 tumor suppressor pathway is activated by defective ribosome synthesis. Ribosomal proteins are released from the nucleolus and block Hdm2 that targets p53 for degradation. However, it remained elusive how abrogation of individual rRNA processing pathways contributes to p53 stabilization. Here we show that selective inhibition of 18S rRNA processing ...

DNA methylation and histone modifications play a central role in the epigenetic regulation of gene expression and cell differentiation. Recently, Np95 (also known as UHRF1 or ICBP90) has been found to interact with Dnmt1 and to bind hemimethylated DNA, indicating together with genetic studies a central role in the maintenance of DNA methylation. Using ...

γ-Secretase is a pivotal intramembrane-cleaving protease complex and important drug target for Alzheimer's disease. The protease not only releases small peptides, such as the amyloid-β peptide, which drives Alzheimer's disease pathogenesis, but also intracellular domains, which can have critical functions in nuclear signaling. Unlike typical aspartyl ...

Protein conformation is critically linked to function and often controlled by interactions with regulatory factors. Here we report the selection of camelid-derived single-domain antibodies (nanobodies) that modulate the conformation and spectral properties of the green fluorescent protein (GFP). One nanobody could reversibly reduce GFP fluorescence by a ...

The heterochromatin-enriched HP1 proteins play a critical role in regulation of transcription. These proteins contain two related domains known as the chromo- and the chromoshadow-domain. The chromo-domain binds histone H3 tails methylated on lysine 9. However, in vivo and in vitro experiments have shown that the affinity of HP1 proteins to ...

The trimeric Sec61/SecY complex is a protein-conducting channel (PCC) for secretory and membrane proteins. Although Sec complexes can form oligomers, it has been suggested that a single copy may serve as an active PCC. We determined subnanometer-resolution cryo–electron microscopy structures of eukaryotic ribosome-Sec61 complexes. In combination with ...

Expression of the Escherichia coli tryptophanase operon depends on ribosome stalling during translation of the upstream TnaC leader peptide, a process for which interactions between the TnaC nascent chain and the ribosomal exit tunnel are critical. We determined a 5.8 angstrom–resolution cryo–electron microscopy and single-particle reconstruction of a ...

Folding and oligomerization of integral membrane proteins frequently depend on specific interactions of transmembrane helices. Interacting amino acids of helix-helix interfaces may form complex motifs and exert different types of molecular forces. Here, a set of strongly self-interacting transmembrane domains (TMDs), as isolated from a combinatorial library, ...

Hypertension and its complications represent leading causes of morbidity and mortality. Although the cause of hypertension is unknown in most patients, genetic factors are recognized as contributing significantly to an individual’s lifetime risk of developing the condition. Here, we investigated the role of the G protein regulator phosducin (Pdc) in ...

Histidine protein kinases and serine, threonine, or tyrosine protein kinases play essential roles in signal transduction in prokaryotes and eukaryotes. A third type of protein kinase, an arginine protein kinase, has been identified. McsB of Bacillus subtilis hosphorylates the heat shock transcriptional regulator CtsR and can be regarded as the founding ...

Closing a gap in the literature, this handbook gathers all the information on single particle tracking and single molecule energy transfer. It covers all aspects of this hot and modern topic, from detecting virus entry to membrane diffusion, and from protein folding using spFRET to coupled dye systems, as well recent achievements in the field. Throughout, ...

The largest subunit of RNA polymerase II (RNAPII) carboxy-terminal heptarepeat domain (CTD) is subject to phosphorylation during initiation and elongation of transcription by RNA polymerase II. Here we study the molecular mechanisms leading to phosphorylation of serine-7 (ser-7) in the human enzyme. Ser-7 becomes phosphorylated before initiation of ...

Assembly and release of human immunodeficiency virus (HIV) occur at the plasma membrane of infected cells and are driven by the Gag polyprotein. Previous studies analyzed viral morphogenesis using biochemical methods and static images, while dynamic and kinetic information has been lacking until very recently. Using a combination of wide-field and total ...

Eukaryotic proteins and in particular cell-surface proteins are frequently glycosylated, which has fueled the interest of chemist to develop new methods for the synthesis of glycosylated proteins. Protein glycosylation is essential for the proper function of the respective proteins and in the case of glycosylated protein therapeutics, such as ...

Chemical Biology: Photosensitive reaction opens or shuts potassium's flow. Drugs that are modulated by light may have a more controllable cousin on the horizon. Researchers led by Dirk Trauner, a chemist at the University of Munich, have figured out the mechanism by which molecules with the ability to block voltage-gated ion channels can be turned on and ...

The adult mouse subependymal zone (SEZ) harbors neural stem cells that are thought to exclusively generate GABAergic interneurons of the olfactory bulb. We examined the adult generation of glutamatergic juxtaglomerular neurons, which had dendritic arborizations that projected into adjacent glomeruli, identifying them as short-axon cells. Fate mapping ...

DNA replication, similar to other cellular processes, occurs within dynamic macromolecular structures. Any comprehensive understanding ultimately requires quantitative data to establish and test models of genome duplication. We used two different super-resolution light microscopy techniques to directly measure and compare the size and numbers of replication ...

Solutions of butylzinc iodide in tetrahydrofuran, acetonitrile, and N,N-dimethylformamide were analyzed by electrospray ionization mass spectrometry. In all cases, microsolvated butylzinc cations [ZnBu(solvent)n]+, n=1-3, were detected. The parallel observation of the butylzincate anion [ZnBuI2]- suggests that these ions result from disproportionation of ...

The PUR protein family is a distinct and highly conserved class that is characterized by its sequence-specific RNA- and DNA-binding. Its best-studied family member, Pur-α, acts as a transcriptional regulator, as host factor for viral replication, and as cofactor for mRNP localization in dendrites. Pur-α-deficient mice show severe neurologic defects and die ...

Many biopharmaceuticals comprise small proteins that are quickly eliminated from the body. CIPSM PI and board member Arne Skerra and his new spin-off XL-protein GmbH combine such small proteins with a kind of molecular balloon that swells and thus prolongs the half-life of these proteins in the body. The CIPSM spin-off XL-protein GmbH has now started to ...

Selective release of enriched biomolecules from solid support is a desirable goal in proteomic and metabolomic studies. Here we demonstrate that photocleavage of a light-sensitive phenacyl ester bond is a suitable alternative cleavage strategy for the selective release of enriched biomolecules form avidin beads circumventing the disadvantages of ...

CIPSM is very proud Thomas Cremer was honored by the prestigious Schleiden Medal of the German Academy of Sciences Leopoldina for his outstanding scientific work on the topograhy of chromosomes in the nucleus of Eucaryota. German press release: Die Deutsche Akademie der Naturforscher Leopoldina - Nationale Akademie der Wissenschaften würdigt die ...

Most of the essential cellular processes such as polymerisation reactions, gene expression and regulation are governed by mechanical processes. Controlled mechanical investigations of these processes are therefore required in order to take our understanding of molecular biology to the next level. Single-molecule manipulation and force spectroscopy have over ...

The translational apparatus is one of the major targets for antibiotics in the bacterial cell. Antibiotics predominantly interact with the functional centers of the ribosome, namely the messenger RNA (mRNA)-transfer RNA (tRNA) decoding region on the 30S subunit, the peptidyltransferase center on the 50S subunit, or the ribosomal exit tunnel through which the ...

Recent studies have indicated that nuclear protein of 95 kDa (Np95) is essential for maintaining genomic methylation by recruiting DNA methyltransferase (Dnmt) 1 to hemi-methylated sites. Here, we show that Np95 interacts more strongly with regulatory domains of the de novo methyltransferases Dnmt3a and Dnmt3b. To investigate possible functions, we developed ...

The cooperation "Experimental and theoretical methods for dissecting the dynamics of epigenetic gene silencing in living cells" between Prof. Heinrich Leonhardt's and Prof. Gunnar Schotta's CIPSM groups and scientists from the University of Heidelberg will be supported for the next three years by the "New methods in systembiology" program ...

CIPSM-Investigator Jens Michaelis has done it! Jens received a grant of nearly 1,5 million Euros from the European Research Council for the next five years. Jens will explore the understanding of the molecular mechanism of nucleosome remodelling using single-molecule fluorescence resonance energy transfer (FRET). In eukaryotic cells the DNA is packaged ...

The translational apparatus is one of the major targets for antibiotics in the bacterial cell. Antibiotics predominantly interact with the functional centers of the ribosome, namely the messenger RNA (mRNA)-transfer RNA (tRNA) decoding region on the 30S subunit, the peptidyltransferase center on the 50S subunit, or the ribosomal exit tunnel through which the ...

We developed a new method for real-time, three-dimensional tracking of fluorescent particles. The instrument is based on a laser-scanning confocal microscope where the focus of the laser beam is scanned or orbited around the particle. Two confocal pinholes are used to simultaneously monitor regions immediately above and below the particle and a feedback loop ...

An important feature of the cerebral cortex is its layered organization, which is modulated in an area-specific manner. We found that the transcription factor AP2gamma regulates laminar fate in a region-specific manner. Deletion of AP2gamma (also known as Tcfap2c) during development resulted in a specific reduction of upper layer neurons in the occipital ...

Modifications make a difference: An isotope-based mass spectrometry method allows the facile and quantitative analysis of modified tRNA nucleosides in various types of cells. This method could be capable of distinguishing between individual cell lines as well as between healthy tissue and cancer cells.

Yeah, the CIPSM football team won the 2009 ScieKickIn football tournament. We thank its members around Prof. Axel Imhof for their excellent performance and passion.

The natural product syringolin A (SylA) is a potent proteasome inhibitor with promising anticancer activities. To further investigate its potential as a lead structure, selectivity profiling with cell lysates was performed. At therapeutic concentrations, a rhodamine-tagged SylA derivative selectively bound to the 20 S proteasome active sites without ...

The DNA of a cell is continuously exposed to numerous endogenous and exogenous factors. The resulting DNA damage can lead to mutations or cell death. Some of the major DNA lesions are generated by the reaction of alkylating reagents with DNAbases. The alkylated reaction products can arise endogenously from cellular alkylating reagents as Sadenosylmethionine ...

Elimination of redundant synapses and strengthening of the surviving ones are crucial steps in the development of the nervous system. Both processes can be readily followed at the climbing fiber to Purkinje cell synapse in the cerebellum. Shortly after birth, around five equally strong climbing fiber synapses are established. Subsequently, one of these five ...

UV-light irradiation induces the formation of highly mutagenic lesions in DNA, such as cis-syn cyclobutane pyrimidine dimers (CPD photoproducts), pyrimidine(6-4)pyrimidone photoproducts ((6-4) photoproducts) and their Dewar valence isomers ((Dew) photoproducts). Here we describe the synthesis of defined DNA strands containing these lesions by direct ...

Cav1.4 channels are unique among the high voltage-activated (HVA) Ca2+ channel family because they completely lack Ca2+-dependent inactivation (CDI) and display very slow voltage-dependent inactivation (VDI). Both properties are of crucial importance in ribbon synapses of retinal photoreceptors and bipolar cells where sustained Ca2+ influx through Cav1.4 ...

Bacteria sense environmental stimuli and transduce this information to cytoplasmic components of the signal transduction machinery to cope with and to adapt to ever changing conditions. Hence, bacteria are equipped with numerous membrane-integrated proteins responsible for sensing such as histidine kinases, chemoreceptors and ToxR-like proteins. There is ...

The dosage compensation complex (DCC) in Drosophila globally increases transcription from the X chromosome in males to compensate for its monosomy.We discovered a male-specific conformation of the X chromosome that depends on the associations of high-affinity binding sites (HAS) of the DCC. The core DCC subunits MSL1–MSL2 are responsible for this ...

The analysis of stress response systems in microorganisms can reveal molecular strategies for regulatory control and adaptation. In this study, we focused on the Cad module, a subsystem of Escherichia coli’s response to acidic stress that is conditionally activated at low pH only when lysine is available. When expressed, the Cad system counteracts the ...

Acetylation is a well-studied posttranslational modification that has been associated with a broad spectrum of biological processes, notably gene regulation. Many studies have contributed to our knowledge of the enzymology underlying acetylation, including efforts to understand the molecular mechanism of substrate recognition by several acetyltransferases, ...

Many marketed drugs contain fluorine, reflecting its ability to modulate a variety of biological responses. The unique 20S proteasome inhibition profile of fluorosalinosporamide compared to chlorinated anticancer agent salinosporamide A (NPI-0052) is exemplary and relates to each halogen’s leaving group potential. Crystal structures of fluoro-, hydroxy-, and ...

We demonstrate the ability to engineer complex shapes that twist and curve at the nanoscale from DNA. Through programmable self-assembly, strands of DNA are directed to form a custom-shaped bundle of tightly cross-linked double helices, arrayed in parallel to their helical axes. Targeted insertions and deletions of base pairs cause the DNA bundles to ...

Transposons are mobile genetic elements found in the hereditary material of humans and other organisms. They can replicate and the new copies can insert at novel sites in the genome. Because this threatens the whole organism, molecular mechanisms have evolved which can repress transposon activity. CIPSM-Professor Klaus Förstemann of the Gene Center of ...

The increasing emergence of multiresistant bacterial pathogens represents a dramatic global health problem. One major reason for this dilemma is the high selective pressure that is exerted on bacteria by classical antibiotic therapies, leading to a steady increase in the number of strains resistant to the majority of all currently available antibiotic drugs. ...

A new photoinducible single electron donor has been developed, which, when linked to thymidine, is shown to be an efficient ground state reducing agent in DNA; the donor can be activated at wavelengths where standard DNA does not absorb.

alpha-Crystallins are molecular chaperones that protect vertebrate eye lens proteins from detrimental protein aggregation. alpha B-Crystallin, 1 of the 2 alpha-crystallin isoforms, is also associated with myopathies and neuropathological diseases. Despite the importance of alpha-crystallins in protein homeostasis, only little is known about their quaternary ...

Quorum sensing (QS) refers to the ability of bacterial populations to read out the local environment for cell density and to collectively activate gene expression. Vibrio harveyi, one of the best characterized model organisms in QS, was used to address the question how single cells behave within a QS-activated community in a homogeneous environment. Analysis ...

Crystallographic studies of the RNA polymerase II (Pol II) elongation complex (EC) revealed the locations of downstream DNA and the DNA-RNA hybrid, but not the course of the nontemplate DNA strand in the transcription bubble and the upstream DNA duplex. Here we used singlemolecule Fluorescence Resonance Energy Transfer (smFRET) experiments to locate ...

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels comprise a small subfamily of proteins within the superfamily of pore-loop cation channels. In mammals, the HCN channel family comprises four members (HCN1-4) that are expressed in heart and nervous system. The current produced by HCN channels has been known as Ih (or If or Iq). Ih has also ...

Eye-opening represents a turning point in the function of the visual cortex. Before eye-opening, the visual cortex is largely devoid of sensory inputs and neuronal activities are generated intrinsically. After eye-opening, the cortex starts to integrate visual information. Here we used in vivo two-photon calcium imaging to explore the developmental changes ...

The KdpD/KdpE two-component system of Escherichia coli regulates expression of the kdpFABC operon encoding the high affinity K+ transport system KdpFABC. The input domain of KdpD comprises a domain that belongs to the family of universal stress proteins (Usp). It has been previously demonstrated that UspC binds to this domain, resulting in KdpD/KdpE ...

Studies have shown that trans-cis isomerization of retinal is the primary photoreaction in the photocycle of the light-driven proton pump bacteriorhodopsin (BR) from Halobacterium salinarum, as well as in the photocycle of the chloride pump halorhodopsin (HR). The transmembrane proteins HR and BR show extensive structural similarities, but differ in the ...

The import of nuclear-encoded preproteins is necessary to maintain chloroplast function. The recognition and transfer of most precursor proteins across the chloroplast envelopes are facilitated by two membrane-inserted protein complexes, the translocons of the chloroplast outer and inner envelope (Toc and Tic complexes, respectively). Several signals have ...

Post-translational histone modifications have essential roles in controlling nuclear processes; however, the specific mechanisms regulating these modifications and their combinatorial activities remain elusive. Cyclin-dependent kinase 9 (CDK9) regulates gene expression by phosphorylating transcriptional regulatory proteins, including the RNA polymerase ...

The ATPase retinoid acid-inducible gene (RIG)-I senses viral RNA in the cytoplasm of infected cells and subsequently activates cellular antiviral defense mechanisms. RIG-I recognizes molecular structures that discriminate viral from host RNA. Here, we show that RIG-I ligands require base-paired structures in conjunction with a free 5′-triphosphate to trigger ...

We apply mesoporous thin silica films with nanometer-sized pores as drug carriers and incorporate the widely used anticancer drug Doxorubicin. Through single-molecule based measurements, we gain mechanistic insights into the drug diffusion inside the mesoporous film, which governs the drug-delivery at the target-site. Drug dynamics inside the nanopores is ...

Cyclic nucleotide-gated (CNG) channels are ion channels which are activated by the binding of cGMP or cAMP. The channels are important cellular switches which transduce changes in intracellular concentrations of cyclic nucleotides into changes of the membrane potential and the Ca2+ concentration. CNG channels play a central role in the signal transduction ...

Members of the AAA+-ATPase superfamily (ATPases associated with various cellular activities) are found in all kingdoms of life and they are involved in very diverse cellular processes, including protein degradation, membrane fusion or cell division. The Arabidopsis genome encodes approximately 140 different proteins that are putative members of this ...

Chloroplast biogenesis in angiosperm plants requires the light-dependent transition from an etioplast stage. A key factor in this process is NADPH:protochlorophyllide oxidoreductase A (PORA), which catalyzes the light-dependent reduction of protochlorophyllide to chlorophyllide. In a recent study the chloroplast outer envelope channel OEP16 was described to ...

Eukaryotes and most prokaryotes require isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) as biosynthetic precursors of terpenes. Whereas animals generate these essential metabolites via the mevalonate pathway,[1] many human pathogens including Plasmodium falciparum and Mycobacterium tuberculosis are known to use the more recently ...

During differentiation and development cells undergo dramatic morphological, and functional changes without any change in the DNA sequence. The underlying changes of gene expression patterns are established and maintained by epigenetic processes. Early mechanistic insights came from the observation that gene activity and repression states correlate with the ...

Second messenger-induced Ca2+-release from intracellular stores plays a key role in a multitude of physiological processes. In addition to 1,4,5-inositol trisphosphate (IP3), Ca2+, and cyclic ADP ribose (cADPR) that trigger Ca2+-release from the endoplasmatic reticulum (ER), nicotinic acid adenine dinucleotide phosphate (NAADP) has been identified as a ...

Nonsense-mediated decay (NMD) is a eukaryotic quality control mechanism that degrades mRNAs carrying premature stop codons. In mammalian cells, NMD is triggered when UPF2 bound to UPF3 on a downstream exon junction complex interacts with UPF1 bound to a stalled ribosome. We report structural studies on the interaction between the C-terminal region of UPF2 ...

Chromosome shattering has been described as a special form of mitotic catastrophe, which occurs in cells with unrepaired DNA damage. The shattered chromosome phenotype was detected after application of a methanol/acetic acid (MAA) fixation protocol routinely used for the preparation of metaphase spreads. The corresponding phenotype in the living cell and the ...

CIPSM sponsors the "Alpenforum 2009" which will rock Oberammergau from 26.-28. of June 2009! The event is dedicated to straighten contacts between young chemists and companies. Be there or be square!

Every cell has to duplicate its entire genome during S-phase of the cell cycle. After replication, the newly synthesized DNA is rapidly assembled into chromatin. The newly assembled chromatin ‘matures’ and adopts a variety of different conformations. This differential packaging of DNA plays an important role for the maintenance of gene expression patterns ...

A prerequisite for antibody secretion and function is their assembly into a defined quaternary structure, composed of two heavy and two light chains for IgG. Unassembled heavy chains are actively retained in the endoplasmic reticulum (ER). Here, we show that the CH1 domain of the heavy chain is intrinsically disordered invitro, which sets it apart from other ...

The localization of mRNAs in subcellular compartments is an efficient way to spatially restrict gene expression. Crystal structures of raver1-vinculin reported by Izard and coworkers now suggest a possible mechanism for mRNA localization during the assembly of focal adhesions.

Neuronal and glial deposition of misfolded, proteolytically processed, polyubiquitinated and abnormally phosphorylated C-terminal fragments (CTFs) of the TAR DNA binding protein-43 (TDP-43) is a pathological hallmark of frontotemporal lobar degeneration with ubiquitin positive inclusions (FTLD-U) and certain cases of amyotrophic lateral sclerosis. We ...

Loss of the of the maintenance methyltransferase xDNMT1 during Xenopus development results in premature transcription and activation of a p53-dependent apoptotic program that accounts for embryo lethality. Here, we show that activation of the apoptotic response is signalled through the methyl-CpG binding protein xMBD4 and the mismatch repair pathway ...

A suitable substitute: All integrin receptors bind their ligands, which contain an aspartate residue, in the metal-ion- dependent adhesion site (MIDAS). So far all attempts to replace the carboxyl group of aspartate with other, pharmacologically favorable isosteric groups have failed. Now it has been shown that a hydroxamic acid group can replace the ...

α5β1-integrins play a key role in angiogenesis, the formation of new vessels in tissues that lack them. By serving as receptors for a variety of extracellular matrix proteins containing an arginine-glycine-aspartic acid (RGD) sequence, these integrins mediate migration of endothelial cells into the basement membrane and regulate their growth, survival, and ...

Photolyases repair cytotoxic and mutagenic UV-induced photolesions in DNA by using an amazing light-dependent repair mechanism. It involves light absorption, electron transfer from an excited reduced and deprotonated FADH− to the flipped-out photolesion, followed by the fragmentation of the photolesions. Cryptochromes are highly related proteins that no ...

In order to promote future scientists the best way we can, CIPSM decided to form an own Graduate School. The Graduate School Life Science Munich (LSM) of the Ludwig-Maximilians-University Munich (LMU) offers an international doctoral program in life sciences covering areas of anthropology, biochemistry and cell biology, ecology, evolution, genetics, ...

The LMU-Harvard Young Scientists’ Forum (YSF) seeks to unite Ph.D. students and postdoctoral fellows from the Harvard University and the Ludwig-Maximilians-Universität (LMU) with core faculty from the two universities to create a framework for an interdisciplinary exchange of ideas. This year’s conference entiteled "From Molecules to Organisms" ...

Science Comment is a new CIPSM spin off dedicated to the advancement of sciences. Feel free to add your comment on publications - be it inspiring thoughts or critics - to propulse the scientific discussion and to enhance future results. On Science Comment publications come under scrutiny. Please use this site to help clarify certain aspects of science. Let's ...

Michael acceptor based natural product derived probes are selective and sensitive chemical tools for the identification and characterization of pathologically relevant enzymes in MRSA.

Posttranslational modifications of the carboxyterminal domain (CTD) of the largest subunit of RNA polymerase II (Pol II) specify a molecular recognition code that is deciphered by proteins involved in RNA biogenesis. The CTD is comprised of a repeating heptapeptide (Y1S2P3T4S5P6S7). Recently, phosphorylation of serine 7 was shown to be important for ...

Click here and keep on scrolling if you care for pictures form the event "CIPSM - Ausgewählter Ort im Land der Ideen 2009" which was held on the 4th of May 2009 in the Deutsche Museum.

Biomolecular reagents that enable the specific molecular recognition of proteins play a crucial role in basic research as well as medicine. Up to now, antibodies (immunoglobulins) have been widely used for this purpose. Their predominant feature is the vast repertoire of antigen-binding sites that arise from a set of 6 hypervariable loops. However, ...

CIPSM ist „Ausgewählter Ort im Land der Ideen“. Damit ist CIPSM Teil der Veranstaltungsreihe „365 Orte im Land der Ideen“, die gemeinsam von der Standortinitiative „Deutschland – Land der Ideen“ und der Deutschen Bank durchgeführt wird. Als „Ausgewählter Ort“ wird CIPSM unter der Schirmherrschaft von Bundespräsident Horst Köhler im Jahr 2009 ...

Archae possess unique biochemical systems quite distinct from the pathways present in eukaryotes and eubacteria. 7,8-Dimethyl-8-hydroxy-5deazaflavin (F0) and F420 are unique deazaflavin-containing coenzyme and methanogenic signature molecules, essential for a variety of biochemical transformations associated with methane biosynthesis and light-dependent DNA ...

Proteins EINtr, NPr and IIANtr form a phosphoryl group transfer chain (Ntr-PTS) working in parallel to the phosphoenolpyruvate:carbohydrate phosphotransferase system (transport-PTS) in Escherichia coli. Recently, it was shown that dephosphorylated IIANtr binds and inhibits TrkA, a low-affinity potassium transporter. Here we report that the Ntr-PTS also ...

Prion disease is characterized by the α→β structural conversion of the cellular prion protein (PrPC) into the misfolded and aggregated “scrapie” (PrPSc) isoform. It has been speculated that methionine (Met) oxidation in PrPC may have a special role in this process, but has not been detailed and assigned individually to the 9 Met residues of full-length, ...

In this Letter, we report the natural products salvianolic acid A, salvianolic acid B, and caftaric acid as inhibitors of the protein–protein interactions mediated by the SH2 domains of the Src-family kinases Src and Lck, two established disease targets. Moreover, we propose a binding mode for the inhibitors based on molecular modeling, which will facilitate ...

Syrbactins, a family of natural products belonging either to the syringolin or glidobactin class, are highly potent proteasome inhibitors. Although sharing similar structural features, they differ in their macrocyclic lactam core structure and exocyclic side chain. These structural variations critically influence inhibitory potency and proteasome subsite ...

Although tremendous progress has been made toward identifying factors that regulate nucleosome structure and positioning, the mechanisms that regulate higher-order chromatin structure remain poorly understood. Recent studies suggest that the ISWI chromatin-remodeling factor plays a key role in this process by promoting the assembly of chromatin containing ...

We show that the nuclear architecture of rod photoreceptor cells differs fundamentally in nocturnal and diurnal mammals. The rods of diurnal retinas possess the conventional architecture found in nearly all eukaryotic cells, with most heterochromatin situated at the nuclear periphery and euchromatin residing toward the nuclear interior. The rods of ...

Astrocytes play many pivotal roles in the adult brain, including their reaction to injury. A hallmark of astrocytes is the contact of their endfeet with the basement membrane surrounding blood vessels, but still relatively little is known about the signaling mediated at the contact site. Here, we examine the role of ß1-integrin at this interface by its ...

We determine the binding mode of a macrocyclic radicicol-like oxime to yeast HSP90 by combining computer simulations and experimental measurements. We sample the macrocyclic scaffold of the unbound ligand by parallel tempering simulations and dock the most populated conformations to yeast HSP90. Docking poses are then evaluated by the use of binding free ...

The transmembrane domains of fusion proteins are known to be functionally important and display an overabundance of helix-destabilizing Ile and Val residues. In an effort to systematically study the relationship of fusogenicity and helix stability, we had previously designed LV-peptides, a low-complexity model system whose hydrophobic core consists of Leu ...

Our aging society is confronted with a dramatic increase of patients suffering from tauopathies, which include Alzheimer disease and certain frontotemporal dementias. These disorders are characterized by typical neuropathological lesions including hyperphosphorylation and subsequent aggregation of TAU protein and neuronal cell death. Currently, no ...

With the development of antibiotic resistant bacterial strains, infectious diseases have become again a life threatening problem. One of the reasons for this dilemma is the limited number and breadth of current therapeutic targets for which several resistance strategies have evolved over time. To identify resistance associated targets and to understand their ...

Magnetofection, gene delivery under the influence of a magnetic field, is a technique to increase transfection efficiency by enforcing gene vector contact with a target cell. Mechanisms of magnetic lipoplex internalization and intracellular details of magnetofection are still unknown. In this study, cellular dynamics of magnetic lipoplexes were examined in ...

HP1 is a major component of chromatin and regulates gene expression through its binding to methylated histone H3. Most eukaryotes express at least three isoforms of HP1 with similar domain architecture. However, despite the common specificity for methylated histone H3, the three HP1 isoforms bind to different regions of the genome. Most of the studies ...

Ischemic cardiomyopathy is one of the main causes of death, which may be prevented by stem cell-based therapies. SDF-1a is the major chemokine attracting stem cells to the heart. Since SDF-1a is cleaved and inactivated by CD26/dipeptidylpeptidase IV (DPP-IV), we established a therapeutic concept— applicable to ischemic disorders in ...

Cyclic nucleotide-regulated cation channels are ion channels whose activation is regulated by the direct binding of cAMP or cGMP to the channel protein. Two structurally related families of channels regulated by cyclic nucleotides have been identified, the cyclic nucleotide-gated channels and the hyperpolarization-activated cyclic nucleotide-gated channels. ...

Nitrile oxides react smoothly and rapidly with norbornene-modified DNA in a copper-free click reaction. The reaction allows high density functionalization of oligodeoxyribonucleotides (ODNs) with a large variety of molecules directly on solid supports and even in synthesizers without the need for an additional catalyst.

Molecular chaperones of the heat shock protein 70 (Hsp70) family play a crucial role in the presentation of exogenous antigenic peptides by antigen-presenting cells (APCs). In a combined biochemical and immuno-logical approach, we characterize the biochemical interaction of tumor-associated peptides with human Hsp70 and show that the strength of this ...

Upon development of radioimmunoassay techniques,[1] radioiodination of peptides and proteins has gradually become an indispensable tool for controlling and monitoring protein and peptide functions in vivo and in vitro, and for studying ligand–receptor interactions, ligand uptake and clearance. In addition, protein iodination has developed into a useful ...

Photoreactions induced by ultraviolet radiation are among the most important external hazards for the integrity of DNA.1 The photolesion with the highest abundance is the [2 + 2] photoaddition of thymine bases adjacent on a DNA strand.2 This photoaddition yields a cyclobutane pyrimidine dimer (CPD). The formation of the CPD lesion has first been ...

2009 is great! If you have time, take 5 minutes to watch CIPSM on silverscreen.

Chemical biology has multiple aims,[1] one of which is the identification of small-molecule modulators for individual functions of as many human proteins as possible. Various approaches towards this goal have been developed, which converge to create a toolbox of small molecules for chemical biologists.[2–4] This endeavor constitutes a long and winding ...

Sequence alignment and database searching are essential tools in biology because a protein’s function can often be inferred from homologous proteins. Standard sequence comparison methods use substitution matrices to find the alignment with the best sum of similarity scores between aligned residues. These similarity scores do not take the local sequence ...

The molecular chaperone heat-shock protein 90 (Hsp90) is one of the most abundant proteins in unstressed eukaryotic cells. Its function is dependent on an exceptionally slow ATPase reaction that involves large conformational changes. To observe these conformational changes and to understand their interplay with the ATPase function, we developed a ...

Intramembrane proteolysis is now widely recognized as an important physiological pathway required for reverse signaling and membrane protein degradation. Aspartyl intramembrane cleaving proteases of the GXGD-type play an important regulatory role in health and disease. Besides -secretase/presenilin, signal peptide peptidase (SPP) and SPP-like (SPPL) ...

The peripheral antenna complex (LH2) of wild type and mutants of the LH2 α-subunit of purple bacterium Rhodobacter sphaeroides are investigated by transient and stationary absorption spectroscopy. The time for energy transfer from the bacteriochlorophyll (BChl) molecules B800 to B850 is found to depend on the mutation and varies between 0.7 ps for wild type ...

The molecular chaperone heat-shock protein 90 (Hsp90) couples ATP hydrolysis to conformational changes driving a reaction cycle that is required for substrate activation. Recent structural analysis provided snapshots of the open and closed states of Hsp90, which mark the starting and end points of these changes. Using fluorescence resonance energy transfer ...

RIG-I is a cytosolic multi-domain protein that detects viral RNA and elicits an antiviral immune response. Two N-terminal caspase activation and recruitment domains (CARDs) transmit the signal and the regulatory domain prevents signaling in the absence of viral RNA. 5’-triphosphate and double stranded (ds) RNA are two molecular patterns that enable RIG-I to ...

For the compact Drosophila genome, several factors mediating insulator function, such as su(Hw) and dCTCF, have been identified. Recent analyses showed that both these insulator-binding factors are functionally dependent on the same cofactor, CP190. Here we analysed genome-wide binding of CP190 and dCTCF. CP190 binding was detected at CTCF, su(Hw) and GAF ...

Human lipocalin 2 (Lcn2), also known as neutrophil gelatinase-associated lipocalin (NGAL), which naturally scavenges bacterial ferric siderophores, has been engineered to specifically bind rare-earth and related metal ions as chelate complexes with [(R)-2-amino-3-(4-aminophenyl)propyl]-trans-(S,S)-cyclohexane-1,2-diaminepentaacetic acid ...

Many metabolic processes essential for plant viability take place in mitochondria. Therefore, mitochondrial function has to be carefully balanced in accordance with the developmental stage and metabolic requirements of the cell. One way to adapt organellar function is the alteration of protein composition. Since most mitochondrial proteins are nuclear ...

The cytidine analogue 5-fluoro-2-deoxycytidine (dCF) is a mechanism-based inhibitor of DNA methyltransferases. We report the synthesis of short 18-mer dsDNA oligomers containing a triple-hemimethylated CpG motive as a recognition sequence for the human methyltransferase Dnmt1. The DNA strands carry within these CpG islands dCF building blocks that function ...

The Tudor-SN protein (p100, SND1) has been implicated in a variety of cellular processes, such as transcription, processing of edited double-stranded RNA, and splicing regulation. Molecular details of these functions are not yet understood. Tudor domains have previously been shown to bind methylated ligands, such as methylated lysines and arginines. It has ...

A wavelength and solvent dependent study of a photochromic indolylfulgide is presented. The ring-closure reaction is characterized using stationary and time-resolved spectroscopy with femtosecond time resolution. After excitation into the first excited singlet state (S1) the photoprocesses proceed on ultrafast timescales (0.3–0.45 ps) in both polar and ...

With decreasing efficiency of antibiotic therapies against hospital- and community-acquired bacterial pathogens, the treatment of infectious diseases again represents a tremendous challenge for medicinal research. This challenge seems to be particularly difficult if one considers the sophisticated resistance strategies, which are effective against almost ...

RIG-I and MDA5 sense cytoplasmic viral RNA and set-off a signal transduction cascade, leading to antiviral innate immune response. The third RIG-Ilike receptor, LGP2, differentially regulates RIG-Iand MDA5-dependent RNA sensing in an unknown manner. All three receptors possess a C-terminal regulatory domain (RD), which in the case of RIG-I senses the viral ...

We herein present a novel platform of well-controlled ordered and disordered nanopatterns positioned with a cyclic peptide of arginine-glycine-aspartic acid (RGD) on a bioinert poly(ethylene glycol) background, to study whether the nanoscopic order of spatial patterning of the integrin-specific ligands influences osteoblast adhesion. This is the first time ...

In order to adapt to ever changing environmental conditions, bacteria sense environmental stimuli, and convert them into signals that are transduced intracellularly. Several mechanisms have evolved by which receptors transmit signals across the cytoplasmic membrane. Stimulus perception may trigger receptor dimerization and/or conformational changes. Another ...

Protein import into peroxisomes depends on a complex and dynamic network of protein–protein interactions. Pex14 is a central component of the peroxisomal import machinery and binds the soluble receptors Pex5 and Pex19, which have important function in the assembly of peroxisome matrix and membrane, respectively. We show that the N-terminal domain of Pex14, ...

Selective base pairing of the four canonical nucleobases is fundamental for the integrity of the genetic system. Information loss associated with DNA damage is a constant challenge and in response, organisms have evolved specialized defence systems consisting of DNA repair and lesion tolerance. DNA repair requires the action of different lesion recognition ...

Polo-like kinases (Plks) are a conserved family of serine/threonine kinases.[1, 2] The family member Plk1 is a key regulator of mitosis[1, 2] and has been identified as a negative prognostic marker for tumor patients.[3, 4] The widespread recognition of Plk1 as a therapeutic target for the treatment of human tumors has triggered numerous drug discovery ...

Mammalian cells are constantly threatened by multiple types ofDNAlesions arising from various sources like irradiation, environmental agents, replication errors or by-products of the normal cellular metabolism. If not readily detected and repaired these lesions can lead to cell death or to the transformation of cells giving rise to life-threatening diseases ...

Hyperpolarization-activated and cyclic nucleotide-gated (HCN) channels belong to the superfamily of voltage-gated pore loop channels. HCN channels are unique among vertebrate voltage-gated ion channels, in that they have a reverse voltage-dependence that leads to activation upon hyperpolarization. In addition, voltage-dependent opening of these channels is ...

Adult neuronal precursors retain the remarkable capacity to migrate long distances from the posterior (subventricular zone) to the most anterior [olfactory bulb (OB)] parts of the brain. The knowledge about the mechanisms that keep neuronal precursors in the migratory stream and organize this long-distance migration is incomplete. Here we show that blood ...

Single-molecule force spectroscopy allows superb mechanical control of protein conformation. We used a custom-built low-drift atomic force microscope to observe mechanically induced conformational equilibrium fluctuations of single molecules of the eukaryotic calcium-dependent signal transducer calmodulin (CaM). From this data, the ligand dependence of the ...

DNA methylation is a major epigenetic modification and plays a crucial role in the regulation of gene expression. Within the family of DNA methyltransferases (Dnmts), Dnmt3a and 3b establish methylation marks during early development, while Dnmt1 maintains methylation patterns after DNA replication. The maintenance function of Dnmt1 is regulated by its large ...

Recent advances have led to insights into the structure of the bacterial ribosome, but little is knownabout the 3D organization of ribosomes in the context of translating polysomes. We employed cryoelectron tomography and a template-matching approach to map 70S ribosomes in vitrified bacterial translation extracts and in lysates of active E. coli ...

Small regulatory RNAs including small interfering RNAs (siRNAs) and microRNAs (miRNAs) guide Argonaute (Ago) proteins to specific target RNAs leading tomRNA destabilization or translational repression. Here, we report the identification of Importin 8 (Imp8) as a component of miRNA-guided regulatory pathways. We show that Imp8 interacts with Ago proteins and ...

γ-Secretase is a unique intramembrane-cleaving protease complex, which cleaves the Alzheimer′s disease-associated b-amyloid precursor protein (APP) and a number of other type I membrane proteins. Human γ-secretase consists of the catalytic subunit presenilin (PS) (PS1 or PS2), the substrate receptor nicastrin, APH-1 (APH-1a or APH-1b), and PEN-2. To ...

The backbone structure is determined by site-directed spin labeling, double electron electron resonance measurements of distances, and modeling in terms of a helix-loop-helix construct for a transmembrane domain that is supposed to line the translocation pathway in the 54.3 kDa Naþ/proline symporter PutP of Escherichia coli. The conformational distribution ...

DNA replication across blocking lesions occurs by translesion DNA synthesis (TLS), involving a multitude of mutagenic DNA polymerases that operate to protect the mammalian genome. Using a quantitative TLS assay, we identified three main classes of TLS in human cells: two rapid and error-free, and the third slow and error-prone. A single gene, REV3L, encoding ...

The dynamics of the ring-closure reaction of three different bis(thiophen-3-yl)maleimides are investigated using ultrafast spectroscopy in the visible range. The structures of the molecules differ with respect to substitution of the thiophene ring and the maleimide. The experiments reveal reaction kinetics which point to the population of an excited ...

Coupling fast triplet–triplet energy transfer (TTET) between xanthone and naphthylalanine to the helix–coil equilibrium in alanine-based peptides allowed the observation of local equilibrium fluctuations in α-helices on the nanoseconds to microseconds time scale. The experiments revealed faster helix unfolding in the terminal regions compared with the ...

We present a simple, non-radioactive assay for DNA methyltransferase activity and DNA binding. As most proteins are studied as GFP fusions in living cells, we used a GFP binding nanobody coupled to agarose beads (GFP nanotrap) for rapid one-step purification. Immobilized GFP fusion proteins were subsequently incubated with different fluorescently labeled ...

Tetrahydrofuran solutions of the products formed in LiCl-mediated zinc insertion reactions into various organic halides RHal were analyzed by anion-mode electrospray ionization (ESI) mass spectrometry. In all cases, organozincate anions were observed. The reactions with RHal, Hal ) Br and I, yielded predominantly mononuclear complexes, such as ZnRHal2 - ...

Four newly synthesized Hemithioindigo-based peptide-switches with changing meta/para-substitution-pattern within the stilbene-part of the molecule are characterized with time-resolved absorption spectroscopy. The different substances undergo a light-induced Z/E-isomerization: the reaction proceeds on a picosecond timescale with time constants <50 ps for ...

Structural investigations are frequently hindered by difficulties in obtaining diffracting crystals of the target protein. Here, we report the crystallization and structure solution of the U2AF homology motif (UHM) domain of splicing factor Puf60 fused to Escherichia coli thioredoxin A. Both modules make extensive crystallographic contacts, contributing to a ...

Electroretinography (ERG) is an established diagnostic technique in clinical ophthalmology and provides objective information about retinal function. This technique is also applied in basic research, where animal models of hereditary retinopathies have significantly contributed to our understanding of the composition of ERG responses in general and how ...

The photophysical properties of three new water-soluble terrylenediimide (WS-TDI) derivatives are investigated and their utilization in biological experiments is demonstrated. Each of these dyes can be excited in the far red region of the visible spectrum, making them good candidates for in-vivo studies. Single-molecule techniques characterize their ...

Electrospray ionization (ESI) of mixtures of organolithium compounds and zinc chloride in tetrahydrofuran produced manifold mono- and polynuclear organozincate anions. Formation of the latter is strongly favored by the incorporation of chloride ligands, which apparently adopt bridging binding modes. Analysis of LinBu/ZnCl2 solutions at different ...

y-Secretase is an intramembrane cleaving aspartyl protease complex intimately implicated in Alzheimer disease pathogenesis. The protease is composed of the catalytic subunit presenilin (PS1 or PS2), the substrate receptor nicastrin (NCT), and two additional subunits, APH-1 (APH-1a, as long and short splice forms (APH-1aL, APH-1aS), or APH-1b) and PEN-2. ...

DNA photolyases and cryptochromes (cry) form a family of flavoproteins that use light energy in the blue/UV-A region for the repair of UV-induced DNA lesions or for signaling, respectively. Very recently, it was shown that members of the DASH cryptochrome subclade repair specifically cyclobutane pyrimidine dimers (CPDs) in UV-damaged single-stranded DNA. ...

Dosage compensation in male Drosophila relies on the X chromosome–specific recruitment of a chromatin-modifying machinery, the dosage compensation complex (DCC). The principles that assure selective targeting of the DCC are unknown. According to a prevalent model, X chromosome targeting is initiated by recruitment of the DCC core components, MSL1 and MSL2, ...

The sensor kinase KdpD and the response regulator KdpE control induction of the kdpFABC operon encoding the high-affinity K+-transport system KdpFABC in response to K+ limitation or salt stress. Under K+ limiting conditions the Kdp system restores the intracellular K+ concentration, while in response to salt stress K+ is accumulated far above the normal ...

The “seventeen kilodalton protein” (Skp) is a predominant periplasmic chaperone of Escherichia coli, which is involved in the biogenesis of abundant outer membrane proteins (OMPs) such as OmpA, PhoE, and LamB. In this study the substrate profile of Skp was investigated in a proteomics approach. Skp was overexpressed in a deficient E. coli strain as a fusion ...

The photoinduced electrocyclic ring-opening of a fluorinated indolylfulgide is investigated by stationary and ultrafast spectroscopy in the UV/vis spectral range. Photoreactions, initiated by optical excitation into the S1 (570 nm) and SN (340 nm) absorption band of the closed isomer, lead to considerable differences in reaction dynamics and quantum yields. ...

The multifunctional nuclear protein positive cofactor 4 (PC4) is involved in various cellular processes including transcription, replication, and chromatin organization. Recently, PC4 has been identifi ed as a suppressor of oxidative mutagenesis in Escherichia coli and Saccharomyces cerevisiae . To investigate a potential role of PC4 in mammalian DNA repair, ...

We have studied the initial phase of cell adhesion as a function of the lateral organization of individual integrin molecules with single-cell force microscopy. Nanostructures, consisting of hexagonally ordered gold dots, were prepared with diblock-copolymer micelle lithography and functionalized with arginine- glycine-aspartate peptides, thus defining ...

Silver-plated DNA: The deposit of a thin metal layer on biomolecules, such as DNA, requires the formation of small, magic-sized metal nuclei. Through the careful design of a reducing chemical functionality in the form of a dialdehyde, the nucleation process and thus the metallization step can be controlled.

YEAH! CIPSM ist „Ausgewählter Ort im Land der Ideen“. Damit ist CIPSM Teil der Veranstaltungsreihe „365 Orte im Land der Ideen“, die gemeinsam von der Standortinitiative „Deutschland – Land der Ideen“ und der Deutschen Bank durchgeführt wird. Als „Ausgewählter Ort“ wird CIPSM unter der Schirmherrschaft von Bundespräsident Horst Köhler im Jahr 2009 ...

Small noncoding RNAs function in concert with Argonaute (Ago) proteins to regulate gene expression at the level of transcription, mRNA stability, or translation. Ago proteins bind small RNAs and form the core of silencing complexes. Here,wereport the analysis of small RNAs associated with human Ago1 and Ago2 revealed by immunoprecipitation and ...

The maintenance of progenitor cells is a crucial aspect of central nervous system development and maturation, and bHLH transcription factors of the E(Spl) subfamily are involved in this process in all vertebrates studied to date. In the zebrafish embryonic neural plate, a large number of E(Spl) genes (her genes) are at play.We review recent data on this ...

The TIM23 complex is the major translocase of the mitochondrial inner membrane responsible for the import of essentially all matrix proteins and a number of inner membrane proteins. Tim23 and Tim50, two essential proteins of the complex, expose conserved domains into the intermembrane space which interact with each other. Here, we describe in vitro ...

We provide a protocol that describes an explant system that allows the dynamics of motor axons to be imaged. This method is based on nerve–muscle explants prepared from the triangularis sterni muscle of mice, a thin muscle that covers the inside of the thorax. These explants, which can be maintained alive for several hours, contain long stretches of ...

An investigation of the precise interactions between damaged DNA and DNA repair enzymes is required in order to understand the lesion recognition step, which is one of the most fundamental processes in DNA repair. Most recently, photoaffinity labeling approaches have enabled the analysis of even transient protein- DNA interactions. Here we report the ...

Serotonin is a major central nervous modulator of physiology and behavior and plays fundamental roles during development and plasticity of the vertebrate central nervous system (CNS). Understanding the developmental control and functions of serotonergic neurons is therefore an important task. In all vertebrates, prominent serotonergic neurons are found ...

Serotonin is a major central nervous modulator of physiology and behavior and plays fundamental roles during development and plasticity of the vertebrate central nervous system (CNS). Understanding the developmental control and functions of serotonergic neurons is therefore an important task. In all vertebrates, prominent serotonergic neurons are found ...

The activation of well-defined numbers of integrin molecules in predefined areas by adhesion of tissue cells to biofunctionalized micro-nanopatterned surfaces was used to determine the minimum number of activated integrins necessary to stimulate focal adhesion formation. This was realized by combining micellar and conventional e-beam lithography, which ...

Tic110 has been proposed to be a channel-forming protein at the inner envelope of chloroplasts whose function is essential for the import of proteins synthesized in the cytosol. Sequence features and topology determination experiments presently summarized suggest that Tic110 consists of six transmembrane helices. Its topology has been mapped by limited ...

Structural investigations are frequently hindered by difficulties in obtaining diffracting crystals of the target protein. Here, we report the crystallization and structure solution of the U2AF homology motif (UHM) domain of splicing factor Puf60 fused to Escherichia coli thioredoxin A. Both modules make extensive crystallographic contacts, contributing to a ...

The incorporation of proline into cyclic peptides seems to be the most promising way to induce -turn structures. Recently, however, it was shown that N-methylated amino acids might be even better suited than proline for introducing turn structures. Another property of proline, the ability to effect cis-peptide bonds, has also been reported for N-methylated ...

Genetic and epigenetic plasticity allows tumors to evade single-targeted treatments. Here we direct Bcl2-specific short interfering RNA (siRNA) with 5¢-triphosphate ends (3p-siRNA) against melanoma. Recognition of 5¢-triphosphate by the cytosolic antiviral helicase retinoic acid–induced protein I (Rig-I, encoded by Ddx58) activated innate immune cells such ...

Genetic and epigenetic plasticity allows tumors to evade single-targeted treatments. Here we direct Bcl2-specific short interfering RNA (siRNA) with 5¢-triphosphate ends (3p-siRNA) against melanoma. Recognition of 5¢-triphosphate by the cytosolic antiviral helicase retinoic acid–induced protein I (Rig-I, encoded by Ddx58) activated innate immune cells such ...

Viscotoxins are small cationic proteins found in European mistletoe Viscum album. They are highly toxic towards phytopathogenic fungi and cancer cells. Heterologous expression of viscotoxins would broaden the spectrum of methods to be applied for better understanding of their structure and function and satisfy possible biopharmaceutical needs. Here, we ...

Photochemical solar energy conversion is considered as an alternative of clean energy. For future light converting nano-machines photosynthetic reaction centers are used as prototypes optimized during evolution. We introduce a reaction scheme for global optimization and simulate the ultrafast charge separation in photochemical energy conversion. Multiple ...

Specific interactions of transmembrane helices play a pivotal role in the folding and oligomerization of integral membrane proteins. The helix–helix interfaces frequently depend on specific amino acid patterns. In this study, a heptad repeat pattern was randomized with all naturally occurring amino acids to uncover novel sequence motifs promoting ...

PUF60 is an essential splicing factor functionally related and homologous to U2AF65. Its C-terminal domain belongs to the family of U2AF (U2 auxiliary factor) homology motifs (UHM), a subgroup of RNA recognition motifs that bind to tryptophan-containing linear peptide motifs (UHM ligand motifs, ULMs) in several nuclear proteins. Here, we show that the Puf60 ...

The metabotropic glutamate receptors type 1 (mGluR1s) are required for a normal function of the mammalian cerebellum. These G-protein-coupled receptors are abundantly expressed in the principle cerebellar cells, namely the Purkinje neurones. Under physiological conditions, mGluR1s are activated during repetitive activity of both afferent glutamatergic ...

UV irradiation of cells gives rise to the formation of cyclobutane pyrimidine dimers (CPD) and so-called (6-4) DNA lesions (Scheme 1). Both lesions are major photoproducts formed in dipyrimidine sequences of double-stranded DNA. Repair of these lesions is essential because of their high mutagenic potential. Particularly important in many organisms are the ...

Dendritic cell (DC) vaccines have emerged as a promising strategy to induce antitumoral cytotoxic T cells for the immunotherapy of cancer. The maturation state of DC is of critical importance for the success of vaccination, but the most effective mode of maturation is still a matter of debate. Whereas immature DC carry the risk of inducing tolerance, ...

Messenger ribonucleoprotein particles (mRNPs) move randomly within nucleoplasm before they exit from the nucleus. To further understand mRNP trafficking, we have studied the intranuclear movement of a specific mRNP, the BR2 mRNP, in salivary gland cells in Chironomus tentans. Their polytene nuclei harbor giant chromosomes separated by vast regions of ...

Histone modifications play an important role in shaping chromatin structure. Here, we describe the use of an in vitro chromatin assembly system from Drosophila embryo extracts to investigate the dynamic changes of histone modifications subsequent to histone deposition. In accordance with what has been observed in vivo, we find a deacetylation of the ...

We have recently identified the archaic cytokine macrophage migration inhibitory factor (MIF) as a non-canonical ligand of the CXC chemokine receptors CXCR2 and CXCR4 in inflammatory and atherogenic cell recruitment. Because its affinity for CXCR2 was particularly high, we hypothesized that MIF may feature structural motives shared by canonical CXCR2 ...

Salinosporamide A (1 (NPI-0052)) is a potent, monochlorinated 20S proteasome inhibitor in clinical trials for the treatment of cancer. To elucidate the role of the chlorine leaving group (LG), we synthesized analogues with a range of LG potentials and determined their IC50 values for inhibition of chymotrypsin-like (CT-L), trypsin-like (T-L), and ...

The structure of the extracellular domain of BMP receptor IA was determined in solution by NMR spectroscopy and compared to its structure when bound to its ligand BMP-2. While most parts of the secondary structure are highly conserved between the bound and unbound forms, large conformational rearrangements can be observed in the b4b5 loop of BMPR-IA, which ...

Our long-term goal is the in vivo expression of intrinsically colored proteins without the need for further posttranslational modification or chemical functionalization by externally added reagents. Biocompatible (Aza)Indoles (Inds)/(Aza)Tryptophans (Trp) as optical probes represent almost ideal isosteric substitutes for natural Trp in cellular ...

The identification of interaction partners in protein complexes is a major goal in cell biology. Here we present a reliable affi nity purifi cation strategy to identify specifi c interactors that combines quantitative SILAC-based mass spectrometry with characterization of common contaminants binding to affi nity matrices (bead proteomes). This strategy can ...

Spiro β-lactone-based proteasome inhibitors were discovered in the context of an asymmetric catalytic total synthesis of the natural product (+)-lactacystin (1). Lactone 4 was found to be a potent inhibitor of the 26S proteasome, while its C-6 epimer (5) displayed weak activity. Crystallographic studies of the two analogues covalently bound to the 20S ...

It is well established that all camelids have unique antibodies circulating in their blood. Unlike antibodies fromother species, these special antibodies are devoid of light chains and are composed of a heavy-chainhomodimer. These so-called heavy-chain antibodies (HCAbs) are expressed after a V–D–J rearrangement and require dedicated constant g-genes. ...

We are very happy that Ulrike Gaul, professor of molecular biology at Rockefeller University New York, will join CIPSM at the Gene Center in 2009 as a newly established CIPSM full professor. This CIPSM professorship for molecular systems biology has been selected for the first Alexander von Humboldt professorship ever to be awarded and comes with an prize ...

Viscotoxins are small cationic proteins found in European mistletoe Viscum album. They are highly toxic towards phytopathogenic fungi and cancer cells. Heterologous expression of viscotoxins would broaden the spectrum of methods to be applied for better understanding of their structure and function and satisfy possible biopharmaceutical needs. Here, we ...

The winners of the second ever Innovation Award of the German BioRegions were announced on the 7th October – Thomas Böttcher together with Dr. Stephan Sieber from CIPSM were awarded joint prizes by Director General from the German Federal Ministry of Education and Research together with Dr. Heinz Bettmann, BioCologne e.V. representing this year’s ...

Very often, the positions of flexible domains within macromolecules as well as within macromolecular complexes cannot be determined by standard structural biology methods. To overcome this problem, we developed a method that uses probabilistic data analysis to combine single-molecule measurements with X-ray crystallography data. The method determines not ...

With the evolution of multidrug resistant bacterial pathogens, infectious diseases pose once again a serious threat to public health. Especially the opportunistic pathogen Staphylococcus aureus has gained importance through the dramatically increasing appearance of methicillin-resistant (MRSA) strains in hospitals and the recent emergence of epidemic ...

Perdeuterated poly(styrene) is introduced as an almost artefactfree and arbitrarily scalable alignment medium for measuring residual dipolar couplings and other anisotropic NMR parameters; the spectral quality achievable in this new medium is demonstrated for HSQC spectra leading to the conformational analysis of staurosporine and homonuclear TOCSY-type ...

In this study we analyze 5-hydroxytryptamine [5-HT]; serotonin) signaling in zebrafish, an increasingly popular vertebrate disease model. We compare and contrast expression of the 5-HT transporter genes slc6a4a and slc6a4b, which identify 5-HT-producing neurons and three novel 5-HT receptors, htr1aa, htr1ab, and htr1bd. slc6a4a and slc6a4b are expressed in ...

Despite their abundance, still little is known about the rather frequent, constantly proliferating progenitors spread throughout the adult mouse brain parenchyma. The majority of these progenitors express the basic-helix-loop-helix transcription factor Olig2, and their number further increases after injury. Here, we examine the progeny of this progenitor ...

Self-assembly guided by molecular recognition has in the past been employed to assemble nanoparticle superstructures like hypercrystals or nanoparticle molecules. An alternative approach, the direct molecule-by-molecule assembly of nanoscale superstructures, was demonstrated recently. Here we present a hybrid approach where we first assemble a pattern of ...

The attachment of labels onto DNA is of utmost importance in many areas of biomedical research and is valuable in the construction of DNA-based functional nanomaterials. The copper(I)-catalyzed Huisgen cycloaddition of azides and alkynes (CuAAC) has recently been added to the repertoire of DNA labeling methods, thus allowing the virtuallyunlimited ...

The neurodegeneration observed in Alzheimer’s disease has been associated with synaptic dismantling and progressive decrease in neuronal activity. We tested this hypothesis in vivo by using two-photon Ca2+ imaging in a mouse model of Alzheimer’s disease. Although a decrease in neuronal activity was seen in 29% of layer 2/3 cortical neurons, 21% of neurons ...

α1-Acid glycoprotein (AGP) is an important drug-binding protein in human plasma and, as an acute-phase protein, it has a strong influence on pharmacokinetics and pharmacodynamics of many pharmaceuticals. We report the crystal structure of the recombinant unglycosylated human AGP at 1.8 Å resolution, which was solved using the new method of ...

With the development of antibiotic-resistant bacterial strains, infectious diseases have become again a life-threatening problem. One of the reasons for this dilemma is the limited number and breadth of current therapeutic targets for which several resistance strategies have evolved over time. To expand the number of addressable enzyme targets and to ...

2008 is great! Ein hochrangiger Wissenschaftler kündigt seine Stelle an der Eliteuniversität Berkeley, weil er lieber an einer deutschen Universität arbeiten möchte - und bringt auch gleich sein ganzes Forschungsteam mit.

Nonproteinogenic amino acids that either occur naturally or are synthesized chemically are becoming important tools in modern drug discovery. In this context, fluorinated amino acids have great potential in the development of novel pharmaceuticals and drugs. To assess whether different fluorinated aromatic amino acid analogues of phenylalanine, tyrosine, ...

RNA interference is a powerful way to study gene function and is frequently combined with microarray analysis. Here we introduce a similar technology at the protein level by simultaneously applying Stable Isotope Labeling by Amino acids in Cell culture (SILAC) and RNA interference (RNAi) to Drosophila SL2 cells. After knockdown of ISWI, an ATP-hydrolyzing ...

Clearance of cellular debris is a critical feature of the developing nervous system, as evidenced by the severe neurological consequences of lysosomal storage diseases in children. An important developmental process, which generates considerable cellular debris, is synapse elimination, in which many axonal branches are pruned. The fate of these pruned ...

More than one hundred and fifty familial Alzheimer's Disease (FAD)-associated missense mutations in presenilins (PS1 and PS2), the catalytic subunit of the γ- secretase complex, cause aberrant Amyloid β-peptide (Aβ) production, by increasing the relative production of the highly amyloidogenic 42 amino acid variant. The molecular mechanism behind this ...

Biological responses to mechanical stress require strain-sensing molecules, whose mechanically induced conformational changes are relayed to signaling cascades mediating changes in cell and tissue properties. In vertebrate muscle, the giant elastic protein titin is involved in strain sensing via its C-terminal kinase domain (TK) at the sarcomeric M-band and ...

The oxazolidinones represent the first new class of antibiotics to enter into clinical usage within the past 30 years, but their binding site and mechanism of action has not been fully characterized. We have determined the crystal structure of the oxazolidinone linezolid bound to the Deinococcus radiodurans 50S ribosomal subunit. Linezolid binds in the A ...

An improved classic Strecker synthesis was elaborated leading to racemic homopropargylglycine (Hpg) in 61% overall yield, while an asymmetric Strecker reaction produced Hpg and the higher homolog 2-aminohept-6-ynoic acid in significantly higher yields and over 80% ee.

in the past decade, the potential of harnessing the ability of nuclear magnetic resonance (NMr) spectroscopy to monitor intermolecular interactions as a tool for drug discovery has been increasingly appreciated in academia and industry. in this Perspective, we highlight some of the major applications of NMR in drug discovery, focusing on hit and lead ...

Neuronal production in the midbrain-hindbrain domain (MH) of the vertebrate embryonic neural tube depends on a progenitor pool called the ‘intervening zone’ (IZ), located at the midbrain-hindbrain boundary. The progressive recruitment of IZ progenitors along the mediolateral (future dorsoventral) axis prefigures the earlier maturation of the MH basal plate. ...

Imprinted genes are important in development and their allelic expression is mediated by imprinting control regions (ICRs). On their DNA-methylated allele, ICRs are marked by trimethylation at H3 Lys 9 (H3K9me3) and H4 Lys 20 (H4K20me3), similar to pericentric heterochromatin. Here, we investigate which histone methyltransferases control this methylation ...

CIPSM Coordinator Horst Kessler wins 2008 Joseph Rudinger Award of the European Peptide Society for his lifetime achievements in peptide chemistry! The Joseph Rudinger Award award is presented „in commemoration of Josef Rudinger’s role in the foundation of the European Peptide Symposia and of the diverse contributions he made to peptide chemistry.” We thank ...

Protein-protein interactions regulate almost all aspects of cellular signaling and aberrant protein-protein interactions have the potential to cause or contribute to human disease. The modulation of these interactions by drug-like molecules would offer previously unavailable opportunities to explore the relevance of pre-selected protein-protein interactions ...

CIPSM-Junior-Group Leader Katja Sträßer has done it! She received a grant of nearly one million Euros from the European Research Council for the next five years. She will explore the understanding of a new aspect of how genetic information is read out by a cell. Many of the individual steps examined separately in the past are coupled with each other to allow ...

EF4 (LepA) is an almost universally conserved translational GTPase in eubacteria. It seems to be essential under environmental stress conditions and has previously been shown to back-translocate the tRNAs on the ribosome, thereby reverting the canonical translocation reaction. In the current work, EF4 was directly visualized in the process of ...

The chloroplast inner envelope membrane contains many integral proteins which differ in the number of a-helices that anchor the protein into the bilayer. For most of these proteins it is not known which pathway they engage to reach their final localisation within the membrane. In yeast mitochondria, two distinct sorting/insertion pathways have been ...

In the mammalian central nervous system, slow synaptic excitation involves the activation of metabotropic glutamate receptors (mGluRs). It has been proposed that C1-type transient receptor potential (TRPC1) channels underlie this synaptic excitation, but our analysis of TRPC1-deficient mice does not support this hypothesis. Here, we show unambiguously that ...

The orphan nuclear receptor NR4A2/Nurr1 is mandatory for the terminal differentiation of mesencephalic dopamine neurons in mammals, but a similar role has remained elusive in the homologous area of the fish brain, the posterior tuberculum. Using loss- and gain-of-function experiments in zebrafish, we show that NR4A2 is indeed responsible for the expression ...

Gene-specific transcription factors are DNA binding proteins that function at the center of the transcriptional machinery to mediate the cellular phenotype in response to extracellular signals. Owing to the lack of enzymatic activities of these intracellular proteins, manipulating the activity of transcription factors by small organic molecules is widely ...

In vivo protein aggregation is strongly linked to the pathogenesis of several incurable celland neurodegenerative diseases including Alzheimer’s disease (AD) and type II diabetes (T2D). In vitro protein aggregation leads to a loss of function and complicates therapeutic application of a number of bioactive proteins or polypeptides such as insulin. Designing ...

What is rheology? Most people are familiar with the basics of rheology from experience with diarrhea or perhaps rheostats. The word rheology was invented in 1929 to name the discipline of a society engaged in the study of how materials deform in response to forces. It was inspired by a quote by Heraclitus: “παντα ρει” translated as “everything flows”. ...

Biomimetic synthesis is an attempt to assemble natural products along biosynthetic lines without recourse to the full enzymatic machinery of nature. We exemplify this with a total synthesis of exiguamine A and the newly isolated natural product exiguamine B. The most noteworthy feature of this work is an oxidative endgame drawing from the complex chemistry ...

We present the implementation of a target function based on Small Angle Scattering data (Gabel et al. Eur Biophys J 35(4):313–327, 2006) into the Crystallography and NMR Systems (CNS) and demonstrate its utility in NMR structure calculations by simultaneous application of small angle scattering (SAS) and residual dipolar coupling (RDC) restraints. The ...

SNARE proteins mediate fusion of intracellular eukaryotic membranes and their -helical transmembrane domains are known to contribute to lipid bilayer mixing. Synthetic transmembrane domain peptides were previously shown to mimic the function of SNARE proteins in that they trigger liposome fusion in a sequence-specific fashion. Here, we performed a detailed ...

Selected residues of transmembrane domain (TM) IX were previously shown to play key roles in ligand binding and transport in members of the Na+/solute symporter family. Using the Na+/proline transporter PutP as a model, a complete Cys scanning mutagenesis of TM IX (positions 324 to 351) was performed here to further investigate the functional significance ...

The spore photoproduct is a unique photolesion, formed in spores upon irradiation with UV light; to investigate the properties of spore photoproduct containing DNA we have synthesized 5S and 5R lesion analogs and incorporated them into DNA.

The zebrafish adult brain contains numerous neural progenitors and is a good model to approach the general mechanisms of adult neural stem cell maintenance and neurogenesis. Here we use this model to test for a correlation between Fgf signaling and cell proliferation in adult progenitor zones. We report expression of Fgf signals (fgf3,4,8a,8b,17b), receptors ...

Gamma-Secretase mediates the final proteolytic cleavage, which liberates Amyloid beta -peptide, the major component of senile plaques in the brain of Alzheimer's disease patients. Therefore gamma -secretase is a prime target for Amyloid beta -peptide lowering therapeutic strategies. gamma -Secretase is a protein complex composed of four different subunits, ...

U2AF homology motifs (UHM) are protein domains that bind peptidic UHM ligand motifs (ULM) and thus form an intricate network of interactions involved in splicing regulation. Here, we report the backbone assignment of the UHM domain of the splicing factor Puf60 as well as 1H, 15N chemical shifts upon binding of the ULM peptides U2AF65 (85–112), SF1 (1–25), ...

Fpg (MutM) is a bacterial base excision repair enzyme that removes the mutagenic and/or replication-block lesions 8-oxoguanine (8-oxodG) and imidazole-ring opened purines (Fapy-derivatives) from DNA. This work shows that Fpg and its eukaryote homologue Ogg1 recognize with high affinity FapydG and bulky N7-benzyl-FapydG (Bz-FapydG). The comparative crystal ...

The potential of peptides as drug candidates is limited by their poor pharmacokinetic properties. Many peptides have a short half-life in vivo and a lack of oral availability. Inspired by the excellent pharmacokinetic profile of cyclosporine, a natural, multiply N-methylated cyclic peptide, we envisioned multiple N-methylation as a promising way to ...

Mitochondria are essential organelles of the eukaryotic cells that are made by expansion and division of pre-existing mitochondria. The majority of their protein constituents are synthesized in the cytosol. They are transported into and put together within the organelle. This complex process is facilitated by several protein translocases. Here we summarize ...

Genetic high-throughput screens have yielded large sets of potential protein-protein interactions now to be verified and further investigated. Here we present a simple assay to directly visualize protein-protein interactions in single living cells. Using a modified lac repressor system, we tethered a fluorescent bait at a chromosomal lac operator array and ...

Actin belongs to the most abundant proteins in eukaryotic cells which harbor usually many conventional actin isoforms as well as actin-related proteins (Arps). To get an overview over the sometimes confusing multitude of actins and Arps, we analyzed the Dictyostelium discoideum actinome in detail and compared it with the genomes from other model organisms. ...

Folding intermediates play a key role in defining protein folding and assembly pathways as well as those of misfolding and aggregation. Yet, due to their transient nature, they are poorly accessible to high-resolution techniques. Here, we made use of the intrinsically slow folding reaction of an antibody domain to characterize its major folding intermediate ...

The precise coordination of the different steps of DNA replication is critical for the maintenance of genome stability. We have probed the mechanisms coupling various components of the replication machinery and their response to polymerase stalling by inhibition of the DNA polymerases in living mammalian cells with aphidicolin. We observed little change ...

In the current issue of Nature Medicine, Faghihi et al. report that a noncoding antisense RNA against β-secretase, also known as BACE1 (β-site amyloid precursor protein (APP)-cleaving enzyme), may contribute to pathogenesis of Alzheimer's disease. BACE1 is a vertebrate-specific enzyme, which, together with presenilin-dependent γ-secretase, cleaves APP to ...

Viral nucleic acids are recognized by specific pattern-recognition receptors of the Toll-like and RIG-I-like receptor families. Synthetic DNA and RNA oligonucleotides can activate the immune system through these receptors and potentiate Ab and CD8 cytotoxic responses to Ags. Systemic application of immunostimulatory oligonucleotides however also results in ...

Cytoplasmic localization and localized translation of messenger RNAs contribute to asymmetrical protein distribution. Recognition of localized mRNAs by RNA-binding proteins can occur in the cytoplasm or, alternatively, co- or post-transcriptionally in the nucleus. In budding yeast, mRNAs destined for localization are bound by the She2 protein before ...

Cell interactions with adhesive surfaces play a vital role in the regulation of cell proliferation, viability, and differentiation, and affect multiple biological processes. Since cell adhesion depends mainly on the nature and density of the adhesive ligand molecules, spatial molecular patterning, which enables the modulation of adhesion receptor clustering, ...

The inhibition of integrin function is a major challenge in medicinal chemistry. Potent ligands are currently in different stages of clinical trials for the antiangiogenic therapy of cancer and agerelated macula degeneration (AMD). The subtype a5b1has recently been drawn into the focus of research because of its genuine role in angiogenesis. In our ...

Fluorescence light microscopy allows multicolor visualization of cellular components with high specificity, but its utility has until recently been constrained by the intrinsic limit of spatial resolution. We applied three-dimensional structured illumination microscopy (3D-SIM) to circumvent this limit and to study the mammalian nucleus. By simultaneously ...

Thioxoamide (thioamide) bonds are nearly isosteric substitutions for amides but have altered hydrogen-bonding and photophysical properties. They are thus well-suited backbone modifications for physicochemical studies on peptides and proteins. The effect of thioxoamides on protein structure and stability has not been subject to detailed experimental ...

H4K20 methylation is a broad chromatin modification that has been linked with diverse epigenetic functions. Several enzymes target H4K20 methylation, consistent with distinct mono-, di-, and trimethylation states controlling different biological outputs. To analyze the roles of H4K20 methylation states, we generated conditional null alleles for the two ...

Imidazolone (dIz) is an abundant, highly mutagenic, and rather unstable DNA lesion that can cause dG!dC transversion mutations. dIz is generated in DNA by a variety of oxidative processes such as type I photooxidation. Herein we report the synthesis of a carbocyclic nucleoside analogue of dIz and of DNA containing this stabilized lesion analogue. The ...

Heterochromatic chromosomal regions undergo large-scale reorganization and progressively aggregate, forming chromocenters. These are dynamic structures that rapidly adapt to various stimuli that influence gene expression patterns, cell cycle progression, and differentiation. Np95-ICBP90 (m- and h-UHRF1) is a histone-binding protein expressed only in ...

Targeted protein degradation is largely performed by the ubiquitin–proteasome pathway, in which substrate proteins are marked by covalently attached ubiquitin chains that mediate recognition by the proteasome. It is currently unclear how the proteasome recognizes its substrates, as the only established ubiquitin receptor intrinsic to the proteasome is ...

Non-canonical amino acids (NAA), as building blocks for peptides and proteins during ribosomal translation, represent a nearly infinite supply of novel functions. The specific selection, activation and tRNA-charging of amino acids by aminoacyl-tRNA synthetases (AARS) in the aminoacylation reaction are essential steps. In most cases, aminoacylation of NAA is ...

The TIM23 (translocase of the mitochondrial inner membrane) complex mediates translocation of preproteins across and their insertion into the mitochondrial inner membrane. How the translocase mediates sorting of preproteins into the two different subcompartments is poorly understood. In particular, it is not clear whether association of two operationally ...

Photorhabdus luminescens is a Gram-negative luminescent enterobacterium and a symbiote to soil nematodes belonging to the species Heterorhabditis bacteriophora. P.luminescens is simultaneously highly pathogenic to insects. This bacterium exhibits a complex life cycle, including one symbiotic stage characterized by colonization of the upper nematode gut, and ...

The serine/threonine kinase polo-like kinase 1 (Plk1) is critically involved in multiple mitotic processes and has been established as an adverse prognostic marker for tumor patients. Plk1 localizes to its substrates and its intracellular anchoring sites via its polo-box domain (PBD), which is unique to the family of polo-like kinases. Therefore, inhibition ...

The serine/threonine kinase Polo-like kinase 1 (Plk1) is overexpressed in many types of human cancers, and has been implicated as an adverse prognostic marker for cancer patients. Plk1 localizes to its intracellular anchoring sites via its polo-box domain (PBD). Here we show that Plk1 can be inhibited by small molecules which interfere with its ...

Intracellular mRNA localization is a common mechanism to achieve asymmetric distributions of proteins. Previous studies have revealed that in a number of cell types, differentmRNA species are localized by the same transport machinery. However, it has been unclear if these individual mRNA species are specifically sorted into separate or common ...

In higher eukaryotes, an unusual C-terminal domain (CTD) is crucial to the function of RNA polymerase II in transcription. The CTD consists of multiple heptapeptide repeats; differences in the number of repeats between organisms and their degree of conservation have intrigued researchers for two decades. Here, we review the evolution of the CTD at the ...

The Oca family is a novel class of autotransporter-adhesins with highest structural similarity in their C-terminal transmembrane region, which supposedly builds a beta-barrel pore in the outer membrane (OM). The prototype of the Oca family is YadA, an adhesin of Yersinia enterocolitica and Yersinia pseudotuberculosis. YadA forms a homotrimeric lollipop-like ...

Forscher der LMU München haben gezeigt, dass sich mit Hilfe von Silbernanoteilchen die Effizienz von Lichtsammelkomplexen enorm steigern lässt. Da es sich hierbei um die zentralen Funktionsträger für das Einsammeln von Licht bei der Photosynthese handelt, könnte diese Entdeckung bedeutsam für die Entwicklung neuartiger Hybrid-Solarzellen sein.

Fluorescent proteins (FP) are widely used as in vivo reporter molecules and are available in multiple colors spanning almost the entire visible light spectrum. Genetically fused to any protein target, FPs offer a powerful tool to study protein localization and dynamics. After the isolation of the prototypical green fluorescent protein (GFP) from the ...

In this article, we follow the history of one of the most abundant, most intensely studied proteins of the eukaryotic cells: actin. We report on hallmarks of its discovery, its structural and functional characterization and localization over time, and point to present days’ knowledge on its position as a member of a large family. We focus on the rather ...

Introducing point mutations into bacterial chromosomes is important for further progress in studies relying on functional genomics, systems- and synthetic biology, and for metabolic engineering. For many investigations, chromosomal systems are required rather than artificial plasmid based systems. Here we describe the introduction of a single point mutation ...

Synaptic integration is modulated by inhibition onto the dendrites of postsynaptic cells. However, presynaptic inhibition at axonal terminals also plays a critical role in the regulation of neurotransmission. In contrast to the development of inhibitory synapses onto dendrites, GABAergic/glycinergic synaptogenesis onto axon terminals has not been widely ...

Peptide bond formation is a fundamental reaction in biology, catalyzed by the ribosomal peptidyl-transferase ribozyme. Although all active-site 23S ribosomal RNA nucleotides are universally conserved, atomic mutagenesis suggests that these nucleobases do not carry functional groups directly involved in peptide bond formation. Instead, a single ribose ...

Signal transducers and activators of transcription (STATs) are a family of transcription factors that are of central importance for cellular signaling and have therefore emerged as attractive target proteins for cell-permeable small molecules. This review outlines the basic concept of STAT signaling, the relevance of individual members of the STAT family ...

With the advances made in surface patterning by micro- and nanotechnology, alternative methods to immobilize biomolecules for different purposes are highly desired. RGD peptides are commonly used to create cell-attractive surfaces for cell-biological and also medical applications. We have developed a fast, one-step method to bind RGD peptides covalently to ...

Antibodies against the neurite outgrowth inhibitor Nogo-A enhance axonal regeneration following spinal cord injury. However, antibodies directed against myelin components can also enhance CNS inflammation. The present study was designed to assess the efficacy of DNA vaccination for generating antibodies against Nogo-A and to study their pathogenic potential ...

A few weeks ago, many of our friends suffered from ‘winter vomiting disease’, a form of gastroenteritis that swept epidemically across Germany. This unpleasant disease is caused by the highly contagious norovirus, a member of the Caliciviridae family of RNA viruses. Caliciviruses are unusual because they dock to sugar residues on the surfaces of cells to ...

We have measured the equilibrium constant for the denaturation transition of the engineered fluorescein-binding lipocalin FluA as a function of pressure and temperature, taking advantage of the fact that the ligand’s fluorescence is almost fully quenched when complexed with the folded protein, but reversibly reappears on denaturation. From the equilibrium ...

Biogenesis of mitochondria depends on the coordinated action of at least six protein translocases present in both mitochondrial membranes. They use different energy sources to drive unidirectional transport of proteins across and into mitochondrial membranes. Here we present an overview on the energetic requirements of different mitochondrial import ...

An optimized protocol for the mild and selective Fukuyama−Mitsunobu reaction was used for mono- and di-N-alkylation on solid support. Thereby, nonfunctionalized aliphatic and aromatic residues are quickly introduced into transiently protected, primary amines of a linear peptide. N-Alkylation can also be used to implement alkyl chains carrying (protected) ...

The thiopeptide class of antibiotics targets the GTPase-associated center (GAC) of the ribosome to inhibit translation factor function. Using X-ray crystallography, we have determined the binding sites of thiostrepton (Thio), nosiheptide (Nosi), and micrococcin (Micro), on the Deinococcus radiodurans large ribosomal subunit. The thiopeptides, by binding ...

Pathogenic bacteria often use effector molecules to increase virulence.vIn most cases, the mode of action of effectors remains unknown. Strains of Pseudomonas syringae pv. syringae (Pss) secrete syringolin A (SylA), a product of a mixed non-ribosomal peptide/polyketide synthetase, in planta1. Here we identify SylA as a virulence factor because a ...

Evolution of multiresistant bacterial strains has meant that infectious diseases once again pose a major threat to public health. Since many antibiotics still target only a limited set of cellular functions, it is a desirable goal to expand the number and breadth of therapeutic targets as well as to gain a deeper understanding of the molecular mechanisms ...

Pieris AG, a biopharmaceutical company developing Anticalins®, a novel class of targeted human protein therapeutics originally developed in the laboratory of CIPSM researcher and Pieris founder Arne Skerra, announced the closing of a Series B financing round led by new investor OrbiMed Advisors LLC, a major global fund dedicated to healthcare, together with Novo Nordisk A/S via its biotech fund. This investment will allow Pieris to advance its lead program, an Anticalin targeting VEGF, into the clinic.

As a first approach to establishing a three-dimensional culture infection model, we studied the growth behavior of the extracellular pathogen Yersinia enterocolitica in three-dimensional collagen gels (3D-CoG). Surprisingly, we observed that plasmidless Y. enterocolitica was motile in the 3D-CoG in contrast to its growth in traditional motility agar at 37°C. ...

The GroEL/GroES chaperonin system mediates protein folding in the bacterial cytosol. Newly synthesized proteins reach GroEL via transfer from upstream chaperones such as DnaK/DnaJ (Hsp70).Here we employed single molecule and ensemble FRET to monitor the conformational transitions of amodel substrate as it proceeds along this chaperone pathway. We find that ...

The importance of protein N terminus sequence composition for cell physiology was recognized more than two decades ago. However, its relevance for chemical protein engineering through an expanded genetic code was demonstrated only very recently. Nature changes the chemistry of the N terminus by posttranslational modifications (PTMs) such as ...

Chromatin serves to package, protect and organize the complex eukaryotic genomes to assure their stable inheritance over many cell generations. At the same time, chromatin must be dynamic to allow continued use of DNA during a cell’s lifetime. One important principle that endows chromatin with flexibility involves ATPdependent ‘remodeling’ factors, which ...

EH domains are protein–protein interaction domains that function in vesicular trafficking and endocytosis. Here, we report the NMR spectral assignments of the high-affinity complex between the second EH domain of Eps15 and a stonin 2 peptide—providing the basis for the characterization of a two-site binding mode.

Heteronuclear residual dipolar one-bond couplings of organic molecules at natural abundance are most easily measured using t2 coupled HSQC spectra. However, inevitably mismatched transfer delays result in phase distortions due to residual dispersive antiphase coherences in such experiments. In this article, slightly modified t2 coupled HSQC experiments with ...

Ectodomain shedding of the amyloid precursor protein (APP) by the two proteases alpha- and ß-secretase is a key regulatory event in the generation of the Alzheimer’s disease amyloid ß peptide (Aß). ß-secretase catalyzes the first step in Aß-generation, whereas a-secretase cleaves within the Aß domain, prevents Aß generation and generates a secreted form of ...

CIPSM-Researchers Heinrich Leonhardt and Ulrich Rothbauer win the Binder-Innovation-Award for their work on the usage of fluorescent nano-antibodies to detect biological structures and processes in living cells.

The preprotein translocon at the inner envelope of chloroplasts (Tic complex) facilitates the import of nuclear-encoded preproteins into the organelle. Seven distinct subunits have been identified so far. For each of those, specific functions have been proposed based on structural prediction or experimental evidence. Three of those subunits possess modules ...

The combination of chromatin structure and the organization of chromosomes in eukaryotic nuclei affects many genome functions. Distinct functional states of genes ranging from ‘highly active’ to ‘silenced’ correlate with particular nucleosome arrangements, histone variants, histone modifications, and interactions of non-histone regulators. Transcription ...

2008 is great! We are very happy that Dirk Trauner will join CIPSM and the Department of Chemistry and Biochemistry of the LMU in August 08 as a newly established CIPSM full professor. Dirk's research centers on the total synthesis of complex natural products and rationally designed molecular probes and their application to biological problems, especially in ...

Hemithioindigo-molecules show a photochromic behavior due to a photo-induced Z/E-isomerization on a picosecond timescale. Changes in the reaction kinetics, caused by polar substituents attached to the thioindigo-moiety, are studied using time-resolved absorption spectroscopy. The experiments reveal that substituents in the thioindigo-part influence the ...

The etiologies of neurodegenerative diseases may be diverse; however, a common pathological denominator is the formation of aberrant protein conformers and the occurrence of pathognomonic proteinaceous deposits. Different approaches coming from neuropathology, genetics, animal modeling and biophysics have established a crucial role of protein misfolding in ...

NMR spectroscopy is a well-established technique for the screening of compound libraries. One of the biggest advantages of NMR spectroscopy in relation to other methods is that it directly detects even weakinteractions between ligand and target molecules, which makes it ideally suited for fragment-based ligand design. In addition, the number of ...

Proline residues affect protein folding and stability via cis/trans isomerization of peptide bonds and by the Cc-exo or -endo puckering of their pyrrolidine rings. Peptide bond conformation as well as puckering propensity can be manipulated by proper choice of ring substituents, e.g. Cc-fluorination. Synthetic chemistry has routinely exploited ...

Full methyl jacket? A complete library of the N-methylated somatostatin cyclopeptidic analogue Veber–Hirschmann peptide cyclo(-PFwKTF-) is performed with the aim of improving its bioavailability. Several analogues from the library were found to bind to the somatostatin receptor in the nanomolar range and one of them shows a significant oral bioavailability ...

The Na+/solute symporter family comprises more than 400 members of pro- and eukaryotic origin. Using the Na+/proline transporter PutP of Escherichia coli as a model, the role of two conserved residues, Ser-340 and Thr-341, is investigated to obtain insights into the mechanism of transport catalyzed by members of this family. Substitution of these amino acids ...

Bayer Schering Pharma (BSP) and CIPSM sign a strategic partnership. The aim of both partners is to initiate a scientific exchange and to carry out joint projects in the field of protein chemistry. With this cooperation the partners wish to simplify and essentially accelerate the transfer of fundamental perceptions into applications, in order to score a ...

Small heat shock proteins (sHsps) are a widespread and diverse class of molecular chaperones. In vivo, sHsps contribute to thermotolerance. Recent evidence suggests that their function in the cellular chaperone network is to maintain protein omeostasis by complexing a variety of non-native proteins. One of the most characteristic features of sHsps is their ...

Remodelling protein nucleic acid interfaces is an important biological task, which is often carried out by nucleic acid stimulated ATPases of the Swi2/Snf2 superfamily. Here we study the mechano-chemical cycle of such an ATPase, namely the catalytic domain of the Sulfolobus solfataricus Rad54 homologue (SsoRad54cd), by means of fluorescence resonance energy ...

Systematic N-methylation of all peptide bonds in the cyclic pentapeptide cyclo(-d-Ala-Ala4-) has been performed yielding 30 different N-methylated derivatives, of which only seven displayed a single conformation on the NMR time scale. The conformation of these differentially N-methylated peptides was recently reported by us (J. Am. Chem. Soc. 2006, 128, 15 ...

The ubiquitin-like SUMO system functions by a cyclic process of modification and demodification, and recent data suggest that the nucleolus is a site of sumoylation–desumoylation cycles.For example, the tumour suppressor ARF stimulates sumoylation of nucleolar proteins. Here, we show that the nucleolar SUMOspecific protease SENP3 is associated with ...

The linker histone H1 binds to the DNA in between adjacent nucleosomes and contributes to chromatin organization and transcriptional control. It is known that H1 carries diverse posttranslational modifications (PTMs), including phosphorylation, lysine methylation and ADP-ribosylation. Their biological functions, however, remain largely unclear. This is in ...

The purpose of this study was to determine the feasibility of a new positron emission tomography (PET) imaging approach using a 18F-labeled αvβ3 integrin antagonist (18F-Galacto-RGD) to monitor the integrin expression after myocardial infarction. METHODS: Male Wister rats were subjected to 20 min of transient left coronary artery occlusion followed by ...

Molecular signals originating at the cell surface are conveyed by a complex system of interconnected signaling pathways to the nucleus. They converge at transcription factors, which in turn regulate the transcription of sets of genes which ultimately determine the cellular phenotype. Whereas enzymes involved in signaling pathways, that is, intracellular ...

Cells respond to a sudden increase in temperature with the transcription of a special set of genes, a phenomenon known as the heat shock response. In the yeast S. cerevisiae, the molecular chaperone Hsp26 is one component of the heat shock response. Hsp26 has the remarkable ability to sense increases in temperature directly and can switch from an inactive ...

Many phenotypic changes of eukaryotic cells due to changes in gene expression depend on alterations in chromatin structure. Processes involved in the alteration of chromatin are diverse and include post-translational modifications of histone proteins, incorporation of specific histone variants, methylation of DNA and ATP-dependent chromatin remodeling. ...

Since the discovery of radial glia as the source of neurons, their heterogeneity in regard to neurogenesis has been described by clonal and time-lapse analysis in vitro. However, the molecular determinants specifying neurogenic radial glia differently from radial glia that mostly self-renew remain ill-defined. Here, we isolated two radial glial subsets that ...

The ATPase RIG-I senses viral RNAs that contain 5`triphosphates in the cytoplasm. It initiates a signaling cascade that activates innate immune response by interferon and cytokine production, providing essential antiviral protection for the host. The mode of RNA 50-triphosphate sensing by RIG-I remains elusive. We show that the C-terminal regulatory domain ...

During the developmental formation of neuronal circuits, redundant synapses are eliminated and persisting synapses strengthened. In the immature cerebellum, climbing fiber–Purkinje cell synapses undergo a pronounced synaptic rewiring, from a multiple innervation around birth to a mono-innervation in adults. An early stage of this process consists in the ...

DNA methyltransferases convert deoxycytidine (dC) nucleobases in DNA into 5-methyldeoxycytidines (dCMe) using the cofactor S-adenosylmethionine (SAM) as the methyl group donor.Methylation of the canonical dC base, particularly in gene promoter regions, induces complex processes, which finally lead to the silencing of the corresponding gene. This epigenetic ...

Signal transducer and activator of transcription 5b (STAT5b) is constitutively activated in many human tumors. Activity of STAT5b requires binding of its Src homology 2 (SH2) domain to certain phosphotyrosine-containing sequences. We have developed a highthroughput assay based on fluorescence polarization that allows screening of chemical libraries for ...

Eps15 homology (EH) domain-containing proteins play a key regulatory role in intracellular membrane trafficking and cell signalling. EH domains serve as interaction platforms for short peptide motifs comprising the residues NPF within natively unstructured regions of accessory proteins. The EH–NPF interactions described thus far are of very low affinity and ...

Single molecule spectroscopy was applied to study the optical properties of native and refolded peridinin–chlorophyll–protein (PCP) complexes. The native system is a trimer with six chlorophyll a (Chl a) molecules, while the refolded one contains two Chl a and resembles structurally and spectroscopically the PCP monomer. The fluorescence emission of single ...

The oxidative DNA lesion, FaPydG rapidly anomerizes to form a mixture of the a and b anomer. To investigate the mutagenic potential of both forms, we prepared stabilized bioisosteric analogues of both configurational isomers and incorporated them into oligonucleotides. These were subsequently used for thermodynamic melting-point studies and for ...

In the mammalian brain, neurogenesis continues only in few regions of the forebrain. The molecular signals governing neurogenesis in these unique neurogenic niches, however, are still ill defined. Here, we show that bone morphogenic protein (BMP)-mediated signaling is active in adult neural stem cells and is crucial to initiate the neurogenic lineage in ...

BACKGROUND: Hemophilia A is currently treated by infusions of the coagulation factor (F) VIII, of which production and purification remain a challenging task. Current purification procedures using immunoaffinity chromatography are cumbersome, expensive, and suffer from the instability of the applied antibody ligands, which elute along with the product and ...

Single-pair fluorescence resonance energy transfer was used to track RNA exiting from RNA polymerase II (Pol II) in elongation complexes. Measuring the distance between the RNA 5´ end and three known locations within the elongation complex allows us determine its position by means of triangulation. RNA leaves the polymerase active center cleft via the ...

The photochemical reaction dynamics of a set of photochromic compounds based on thioindigo and stilbene molecular parts (hemithioindigos, HTI) are presented. Photochemical Z/E isomerization around the central double bond occurs with time constants of 216 ps (Z → E) and 10 ps (E → Z) for a 5-methyl-hemithioindigo. Chemical substitution on the stilbene moiety ...

Synthesis of ribosomal RNA (rRNA) by RNA polymerase (Pol) I is the first step in ribosome biogenesis and a regulatory switch in eukaryotic cell growth. Here we report the 12 A° cryoelectron microscopic structure for the complete 14-subunit yeast Pol I, a homology model for the core enzyme, and the crystal structure of the subcomplex A14/43. In the resulting ...

Ensemble and single-molecule spectroscopy demonstrates that both emission and absorption of peridinin-chlorophyll-protein photosynthetic antennae can be largely enhanced through plasmonic interactions. We find up to 18-fold increase of the chlorophyll fluorescence for complexes placed near a silver metal layer. This enhancement, which leaves no measurable ...

The amoeba Dictyostelium discoideum possesses genes for 13 different kinesins. Here we characterize DdKif3, a member of the Kinesin-1 family. Kinesin-1 motors form homodimers that can move micrometer-long distances on microtubules using the energy derived from ATP hydrolysis. We expressed recombinant motors in Escherichia coli and tested them in different in ...

Loss of function of the RNA helicase maleless (MLE) in Drosophila melanogaster leads to male-specific lethality due to a failure of X chromosome dosage compensation. MLE is presumably involved in incorporating the non-coding roX RNA into the dosage compensation complex (DCC), which is an essential but poorly understood requirement for faithful targeting of ...

In an acidic (pH 5.8) and lysine-rich environment Escherichia coli induces expression of the cadBA operon which encodes CadA, catalysing the decarboxylation of lysine to cadaverine, and CadB, the lysine/cadaverine antiporter. cadBA expression is dependent on CadC, a membrane-integrated transcriptional activator which belongs to the ToxR-like protein family ...

Patters of phosphorylation in a region of RNA polymerase II may constitute a code that controls the recruitment of regulatory factors to control gene expression. Jeffry L. Corden puts CIPSM-researcher Dirk Eick's recent work on transcription in perspective. The development of higher forms of life would appear to have been influenced by RNA polymerase II. ...

RNA polymerase II is distinguished by its large carboxyl-terminal repeat domain (CTD), composed of repeats of the consensus heptapeptide Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. Differential phosphorylation of serine-2 and serine-5 at the 5′ and 3′ regions of genes appears to coordinate the localization of transcription and RNA processing factors to the ...

RNA polymerase II (Pol II) transcribes genes that encode proteins and noncoding small nuclear RNAs (snRNAs). The carboxyl-terminal repeat domain (CTD) of the largest subunit of mammalian RNA Pol II, comprising tandem repeats of the heptapeptide consensus Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7, is required for expression of both gene types. We show that mutation ...

Genome activity and nuclear metabolism clearly depend on accessibility, but it is not known whether and to what extent nuclear structures limit the mobility and access of individual molecules. We used fluorescently labeled streptavidin with a nuclear localization signal as an average-sized, inert protein to probe the nuclear environment. The protein was ...

The Deutsche Forschungsgemeinschaft (German Research Foundation, DFG) announced yesterday the winners of its 2008 Gottfried Wilhelm Leibniz Prize. At its meeting on December 6th 2007, the DFG Joint Committee named CIPSM researcher Dr. Elena Conti as recipient of Germany's most highly endowed research award. Elena will receive 1.25 million euros flexibly over ...

Protein synthesis in the chloroplast is carried out by chloroplast ribosomes (chloro-ribosome) and regulated in a light-dependent manner. Chloroplast or plastid ribosomal proteins (PRPs) generally are larger than their bacterial counterparts, and chloro-ribosomes contain additional plastid-specific ribosomal proteins (PSRPs); however, it is unclear to what ...

Langerin is a type II transmembrane oligosaccharide receptor on Langerhans cells (LCs), a prominent subclass of dendritic cells (DCs) that mediate immune responses in epithelia and play a role in HIV degradation. Its extracellular moiety comprises a neck region with several heptad repeats and an exposed carboxy-terminal calcium-type carbohydrate-recognition ...

Periodic mesoporous materials formed through the cooperative self-assembly of surfactants and framework building blocks can assume a variety of structures1–3, and their widely tuneable properties make them attractive hosts for numerous applications4–7. Because the molecular movement in the pore system is the most important and defining characteristic of ...

The motor protein kinesin ‘walks’ by alternately advancing its two motor structural domains. A cutting-edge, single-molecule fluorescencetechnique reveals further details of this stepping mechanism. Suppose that walking required energy input in the form of, say, one Gummi bear for every step. In what position would you pop the next Gummi bear into your ...

After the complete sequencing of the human genome, one of the next important steps is to decipher the genetic mechanisms which allow cells to switch genes on and off. These processes are fundamental for cellular development. At the chemical level, gene silencing requires the selective methylation of of the C5-position of cytidines in DNA, and ...

LV-peptides were designed as membrane-spanning low-complexity model structures that mimic fusion protein transmembrane domains. These peptides harbor a hydrophobic core sequence that consists of helix-promoting and helix-destabilizing residues at different ratios. Previously, the fusogenicity of these peptides has been shown to increase with the ...

CIPSM-Researcher and Speaker Thomas Carell wins the prestigious Otto-Bayer-Preis 2008 for his work of the understanding of light-triggered DNA-lesions. Thomas is a specialist for the synthesis of DNA-lesions and lesion analogues, and their incorporation into DNA-strands and the investigations of the resulting structures (x-ray, enzymatic digestion) and ...

Nature takes in its spotlight on Germany a glimpse of CIPSM.

RNA polymerase (Pol) II catalyses DNA-dependent RNA synthesis during gene tran- scription. There is, however, evidence that Pol II also possesses RNA-dependent RNA polymerase (RdRP) activity. Pol II can use a homopolymeric RNA template, can extend RNA by several nucleotides in the absence of DNA, and has been implicated in the replication of the RNA genomes ...

"There's great excitement here" “If the show is over in five years, then it was but a flash in the pan”

CISPLATIN HAS BEEN USED to treat cancer for decades. But the drug's detailed molecular interactions with the enzymes it aims to interrupts have remained obscure. In two separate papers, researchers are now reporting the first crystal structures of cisplatin complexed with two key enzymes. "For the first time, we are getting deep insight on what goes on ...

The general transcription factors (GTFs) of eukaryotic RNA polymerase II, in a process facilitated by regulatory and accessory factors, target promoters through synergistic interactions with core elements. The specific binding of the TATA box–binding protein (TBP) to the TATA box has led to the assumption that GTFs recognize promoters directly, producing a ...

The anticancer drug cisplatin forms 1,2-d(GpG) DNA intrastrand cross-links (cisplatin lesions) that stall RNA polymerase II (Pol II) and trigger transcription-coupled DNA repair. Here we present a structure-function analysis of Pol II stalling at a cisplatin lesion in the DNA template. Pol II stalling results from a translocation barrier that prevents ...

Ribosome binding factor A (RbfA) is a bacterial cold shock response protein, required for an efficient processing of the 5' end of the 16S ribosomal RNA (rRNA) during assembly of the small (30S) ribosomal subunit. Here we present a crystal structure of Thermus thermophilus (Tth) RbfA and a three-dimensional cryo-electron microscopic (EM) map of the Tth ...

DNA polymerase eta (Pol eta) is a eukaryotic lesion bypass polymerase that helps organisms to survive exposure to ultraviolet (UV) radiation, and tumor cells to gain resistance against cisplatin-based chemotherapy. It allows cells to replicate across cross-link lesions such as 1,2-d(GpG) cisplatin adducts (Pt-GG) and UVinduced cis–syn thymine dimers. We ...

Site-directed mutagenesis (SDM) of target DNA is an invaluable tool to study protein structure–function relationships. Alanine-scanning mutagenesis has been successfully applied to systematically map functional binding epitopes. Substitution of target amino acids with alanine removes all side chain atoms past the b-carbon and does not introduce unusual ...

Protein synthesis initiates with Met in the cytosol of eukaryotes and formylmethionine (fMet) in prokaryotes and eukaryotic organelles. N-terminal methionine excision (NME) is the major source of N-terminal amino acid diversity in all three life kingdoms. The excision is dictated by the nature and bulkiness of the side-chain of the second amino acid (the ...

Degeneration of dopaminergic neurons in the substantia nigra is characteristic for Parkinson’s disease (PD), the second most common neurodegenerative disorder. Mitochondrial dysfunction is believed to contribute to the etiology of PD. Although most cases are sporadic, recent evidence points to anumberof genes involved in familial variants of PD.Amongthem, a ...

Loss of function mutations in progranulin cause tau-negative frontotemporal lobar degeneration with ubiquitin positive inclusions. A major protein component of these inclusions is TDP-43, which becomes hyperphosphorylated, ubiquitinated and cleaved to generate C-terminal fragments, which apparently translocate from nuclei to the cytoplasm. Most ...

Genome integrity is constantly threatened by DNA lesions arisin from numerous exogenous and endogenous sources. Survival depends on immediate recognition of these lesions and rapid recruitment of repair factors. Using laser microirradiation and live cell microscopy we found that the DNA-damage dependent poly(ADP-ribose) polymerases (PARP) PARP-1 and PARP-2 ...

An unselective cyclic peptide integrin ligand was sequentially N-methylated by a designed approach, where only the externally oriented (solvent exposed) amide bonds were N-methylated. The N-methylation resulted in tremendous enhancement in selectivity among the different integrin receptor subtypes (alpha5beta1, alphavbeta3, and alphaIIbbeta3). ...

In a recent issue of Molecular Cell, Dollins et al. (2007) present the crystal structure of Grp94, which highlights the similarity between Grp94 and Hsp90 and provides insight into the resting state of Grp94 and potentially other Hsp90 family members.

The genome of the hyperthermophile archaeon Pyrococcus furiosus encodes two transcription factor B (TFB) paralogs, one of which (TFB1) has been characterized in transcription initiation. The second TFB (TFB2) is unusual in that it lacks recognizable homology to the archaeal TFB/eukaryotic TFIIB B-finger motif. TFB2 functions poorly in promoter dependent ...

In a recent issue of Molecular Cell, Dollins et al. (2007) present the crystal structure of Grp94, which highlights the similarity between Grp94 and Hsp90 and provides insight into the resting state of Grp94 and potentially other Hsp90 family members.

Presenilin, the catalytic component of the gamma-secretase complex, type IV prepilin peptidases and signal peptide peptidase (SPP) are the founding members of the family of intramembrane cleaving GxGD aspartyl proteases. SPP-like (SPPL) proteases, such as SPPL2a, b, and c and SPPL3 also belong to the GxGD family. In contrast to gamma-secretase, where ...

Peridinin–chlorophyll–protein (PCP), containing differently absorbing chlorophyll derivatives, are good models with which to study energy transfer among monomeric chlorophylls (Chls) by both bulk and singlemolecule spectroscopy. They can be obtained by reconstituting the N-terminal domain of the protein (N-PCP) with peridinin and chlorophyll mixtures. Upon ...

Alzheimer's disease (AD) amyloid beta-peptide (Aß) is generated via proteolytic processing of the ß-amyloid precursor protein (APP) by ß- and gamma-secretase. gamma-Secretase can be blocked by selective inhibitors but can also be modulated by a subset of non-steroidal anti-inflammatory drugs (NSAIDs) including sulindac sulfide. These drugs selectively reduce ...

Green fluorescent proteins (GFPs) and variants thereof are widely used to study protein localization and dynamics. We engineered a specific binder for fluorescent proteins based on a 13-kDa GFP binding fragment derived from a llama single chain antibody. This GFP-binding protein (GBP) can easily be produced in bacteria and coupled to a monovalent matrix. The ...

Human apolipoproteinD(ApoD) occurs in plasma associated with high density lipoprotein. Apart from the involvement in lipid metabolism, its binding activity for progesterone and arachidonic acid plays a role in cancer development and neurological diseases. The crystal structures of freeApoDand its complex with progesterone were determined at 1.8A˚ ...

RNA polymerases (Pol) II and III synthesize eukaryotic mRNAs and tRNAs, respectively. The crystal structure of the 12 subunit Pol II is known, but only limited structural information is available for the 17 subunit Pol III. Using mass spectrometry (MS), we correlated masses of Pol II complexes with the Pol II structure. Analysis of Pol III showed that the ...

Grp94, the Hsp90 paralogue of the endoplasmic reticulum (ER), plays a crucial role in protein secretion. Like cytoplasmic Hsp90, Grp94 is regulated by nucleotide binding to its N-terminal domain. However, the question whether Grp94 hydrolyses ATP was controversial. This sets Grp94 apart from other members of the Hsp90 family where a slow but specific ...

MicroRNAs (miRNAs) control gene expression by regulating mRNA stability and translation. Using cell-free in vitro systems, several labs have recently reported insights into the molecular mechanisms underlying miRNA-guided translational repression (Kiriakidou et al., 2007; Mathonnet et al., 2007; Thermann and Hentze, 2007; Wakiyama et al., 2007). These new ...

A light-switchable peptide is transformed with ultrashort pulses from aß-hairpin to an unfolded hydrophobic cluster and vice versa. The structural changes are monitored by mid-IR probing. Instantaneous normal mode analysis with a Hamiltonian combining density functional theory with molecular mechanics is used to interpret the absorption transients. ...

The import of proteins destined for the intermembrane space of chloroplasts has not been investigated in detail up to now. By investigating energy requirements and time courses, as well as performing competition experiments, we show that the two intermembrane space components Tic22 and MGD1 (E.C. 2.4.1.46) both engage the Toc machinery for crossing the outer ...

Interactions of transmembrane helices play a crucial role in the folding and oligomerisation of integral membrane proteins. In order to uncover novel sequence motifs mediating these interactions, we randomised one face of a transmembrane helix with a set of non-polar or moderately polar amino acids. Those sequences capable of self-interaction upon ...

The main purpose of homonuclear Hartmann–Hahn or TOCSY experiments is the assignment of spin systems based on efficient coherence transfer via scalar couplings. In partially aligned samples, however, magnetization is also transferred via residual dipolar couplings (RDCs) and therefore through space correlations can be observed in COSY and TOCSY experiments ...

Dynamic gene repositioning has emerged as an additional level of epigenetic gene regulation. An early example was the report of a transient, spatial convergence (<2 micro m) of oppositely imprinted regions (‘‘kissing’’), including the Angelman syndrome/Prader– Willi syndrome (AS/PWS) locus and the Beckwith–Wiedemann syndrome locus in human ...

Channels directly gated by cyclic nucleotides (CNG channels) are important cellular switches that mediate influx of Na+ and Ca2+ in response to increases in the intracellular concentration of cAMP and cGMP. In photoreceptors and olfactory receptor neurons, these channels serve as final targets for cGMP and cAMP signaling pathways that are initiated by the ...

The mammalian main olfactory epithelium (MOE) recognizes and transduces olfactory cues through a G protein-coupled, cAMPdependent signaling cascade. Additional chemosensory transduction mechanisms have been suggested but remain controversial. We show that a subset of MOE neurons expressing the orphan receptor guanylyl cyclase GC-D and the cyclic ...

Quantum dot (QD) bioimaging has recently been described as the most exciting new technique to emerge from the collaboration of physicists and biologists. This new molecular imaging technology is based on the selective fluorescent labeling of biological molecules. QDs are fluorescent nanocrystals showing a wide-range absorption spectrum (400–650 nm) and a ...

Transient or partial formation of complexes between biomacromolecules is a general mechanism used to control cellular functions. Several of these complexes escape structure determination by crystallographic means. We developed a new approach for determining the structure of protein dimers in the native environment (e.g., in the membrane) with high resolution ...

Direct observation of the folding of a single polypeptide chain can provide important information about the thermodynamic states populated along its folding pathway. In this study we present a lock-in force spectroscopy technique that improves resolution of AFM force spectroscopy to 400 fN. Using this technique we show that immunoglobulin domain 4 from ...

Hemophilia A, one of the most severe bleeding disorders, results from an inherited deficiency of factor VIII (FVIII) function. Treatment by injection of FVIII has been a common procedure for decades. Nevertheless, the production and purification of FVIII remains a challenging task. Current procedures using immunoaffinity chromatography are expensive and ...

gamma-Secretase is an intramembrane aspartyl protease complex that cleaves type I integral membrane proteins, including the amyloid beta-protein precursor (APP) and the Notch receptor, and is composed of presenilin, Pen-2, nicastrin and Aph-1. Although all four of these membrane proteins are essential for assembly and activity, the stoichiometry of the ...

Infrared absorption experiments on light-triggered azobenzene peptides have been performed below and above the freezing point of the solvent dimethyl sulfoxide (DMSO). Even 20 K below the freezing point, illumination of the azobenzene chromophore resulted in IR absorption changes indicative of light-induced structural rearrangements of the peptide. In ...

How does RNA polymerase II cooperate with initiation factors to locate transcription start sites throughout the genome? A new cross-linking approach reveals previously unknown initiation factor–binding sites on the polymerase surface. The resulting model of the transcription initiation complex suggests that initiation factors cooperate above and inside the ...

Small interfering RNAs (siRNAs) and microRNAs (miRNAs) guide distinct classes of RNA-induced silencing complexes (RICSs) to repress mRNA expression in biological processes ranging from development to antiviral defense. In Drosophilia, separate but conceptually similar endonucleolytic pathways produce siRNAs and miRNAs. Here, we show that despite their ...

During gene transcription, RNA polymerase (Pol) II encounters obstacles, including lesions in the DNA template. Here, we review a recent structure–function analysis of Pol II transcribing DNA with a bulky photo-lesion in the template strand. The study provided the molecular basis for recognition of a damaged DNA by Pol II, which is the first step in ...

In addition to miRNAs and siRNAs, a third small RNA silencing system has been uncovered that prevents the spreading of selfish genetic elements. Production of the Piwi-associated RNAs (piRNAs), which mediate the silencing activity in this pathway, is initiated at a few master control regions within the genome. The nature of the primary piRNA-generating ...

Gamma-Secretase is involved in the production of amyloid beta-peptide, which is the principle component of amyloid plaques in the brains of patients with Alzheimer disease. Gamma-Secretase is a complex composed of presenilin (PS), nicastrin, anterior pharynx-defective phenotype 1 (Aph1) and PS enhancer 2 (Pen2). We previously proposed a mechanism of complex ...

The molecule HTI, which combines hemithioindigo and hemistilbene molecular parts, allows reversible switching between two isomeric states. Photochromic behaviour of the HTI molecule is observed by irradiation with UV/Vis light. The photochemical reaction, a Z/E isomerization around the central double bond connecting the two molecular parts, is investigated ...

Fibronectin (FN) is secreted as a disulfi de-bonded FN dimer. Each subunit contains three types of repeating modules: FN-I, FN-II, and FN-III. The interactions of α5β1 or αv integrins with the RGD motif of FN-III repeat 10 (FN-III10) are considered an essential step in the assembly of FN fi brils. To test this hypothesis in vivo, we replaced the RGD motif ...

The U2AF-homology motif (UHM) mediates protein-protein interactions between factors involved in constitutive RNA splicing. Here we report that the splicing factor SPF45 regulates alternative splicing of the apoptosis regulatory gene FAS (also called CD95). The SPF45 UHM is necessary for this activity and binds UHM-ligand motifs (ULMs) present in the 3' ...

To reveal the mechanism of processive strand separation by superfamily-2 (SF2) 3'-5' helicases, we determined apo and DNA-bound crystal structures of archaeal Hel308, a helicase that unwinds lagging strands and is related to human DNA polymerase h. Our structure captures the duplex-unwinding reaction, shows that initial strand separation does not require ...

During gene transcription, RNA polymerase (Pol) II encounters obstacles, including lesions in the DNA template. Here, we review a recent structure–function analysis of Pol II transcribing DNA with a bulky photo-lesion in the template strand. The study provided the molecular basis for recognition of a damaged DNA by Pol II, which is the first step in ...

The Strep-tag II is an eight-residue minimal peptide sequence (Trp-Ser-His-Pro-Gln-Phe-Glu-Lys) that exhibits intrinsic affinity toward streptavidin and can be fused to recombinant proteins in various fashions. We describe a protocol that enables quick and mild purification of corresponding Strep-tag II fusion proteins—including their complexes with ...

The photochromicity of fulgimides rests on the existence of open (E) and closed ring (C) isomers. As predicted by the Woodward-Hoffmann rules both isomers can photochemically be interconverted. This interconversion has been studied by femtosecond fluorescence and transient absorption spectroscopy. For either direction (E toC cyclization and C to E ...

A novel type of non-linear Raman microscopy, femtosecond stimulated Raman microscopy (FSRM), is introduced. It employs femtosecond white light pulses and intense picosecond pulses which are derived from a femtosecond laser/amplifier system. The pulses are coupled into a microscope set-up and induce a stimulated Raman process at the focus. The Raman ...

The bacteriophage Phi29 uses a molecular motor to drive its genome into a preformed protein capsid. The central part of this molecular motor is formed by a ring of twelve proteins called the connector. Symmetry and structural arguments have let to the so called rotation hypothesis, i.e. a rotation of the connector is used to drive the DNA into the capsid, ...

Cells use transcription-coupled repair (TCR) to efficiently eliminate DNA lesions such as UV-induced cyclobutane pyrimidine dimers (CPDs). Here we present the structure-based mechanism for the first step in eukaryotic TCR, CPD-induced stalling of RNA polymerase (Pol) II. A CPD in the transcribed strand slowly passes a translocation barrier, and enters the ...

The assembly of the gamma-secretase complex, an Alzheimer disease-related protease required for ß-amyloid generation, is tightly regulated and predominantly limited by the stoichiometrical availability of its components. We have identified a novel endoplasmic reticulum-located protein complex that is regulatedin a similar fashion. It contains the recently ...

CIPSM researcher and LMU professor Patrick Cramer wins this year the prestigious award of the Philip Morris Foundation. Patrick is honored for his research of the enzyme RNA-polymerase II, which is essential for translation of genetic information in proteins.

The Deutsche Forschungsgemeinschaft (German Research Foundation, DFG) has announced the winners of its 2007 Gottfried Wilhelm Leibniz Prize. At its meeting on 7 December 2006, the DFG Joint Committee named CIPSM researcher and LMU Professor Magdalena Götz as recipient of Germany's most highly endowed research award.