Christian Griesinger

Max Planck Institute for Biophysical Chemistry, Göttingen, Lower Saxony, Germany

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Publications (371)1728.97 Total impact

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    ABSTRACT: The eIF4E-binding protein 1 (4EBP1) has long been known to be completely unstructured without any secondary structures, which contributed significantly to the proposal of the induced fit mechanism for target binding of intrinsically disordered proteins. We show here that 4EBP1 is not completely unstructured, but contains a pre-structured helix.
    Molecular BioSystems 11/2014; · 3.35 Impact Factor
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    ABSTRACT: The voltage-dependent anion channel (VDAC) is the most abundant protein of the outer mitochondrial membrane and constitutes the major pathway for the transport of ADP, ATP, and other metabolites. In this multidisciplinary study we combined solid-state NMR, electrophysiology, and molecular dynamics simulations, to study the structure of the human VDAC isoform 2 in a lipid bilayer environment. We find that the structure of hVDAC2 is similar to the structure of hVDAC1, in line with recent investigations on zfVDAC2. However, hVDAC2 appears to exhibit an increased conformational heterogeneity compared to hVDAC1 which is reflected in broader solid-state NMR spectra and less defined electrophysiological profiles.
    Journal of Biomolecular NMR 11/2014; · 2.85 Impact Factor
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    ABSTRACT: Motions play a vital role in the functions of many proteins. Discrete conformational transitions to excited states, happening on timescales of hundreds of microseconds, have been extensively characterized. On the other hand, the dynamics of the ground state are widely unexplored. Newly developed high-power relaxation dispersion experiments allow the detection of motions up to a one-digit microsecond timescale. These experiments showed that side chains in the hydrophobic core as well as at protein-protein interaction surfaces of both ubiquitin and the third immunoglobulin binding domain of protein G move on the microsecond timescale. Both proteins exhibit plasticity to this microsecond motion through redistribution of the populations of their side-chain rotamers, which interconvert on the picosecond to nanosecond timescale, making it likely that this "population shuffling" process is a general mechanism.
    Angewandte Chemie International Edition in English 11/2014; · 13.45 Impact Factor
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    ABSTRACT: Intrinsically disordered proteins (IDPs) are involved in a wide variety of physiological and pathological processes and are best described by ensembles of rapidly interconverting conformers. Using fast field cycling relaxation measurements we here show that the IDP alpha-synuclein as well as a variety of other IDPs undergoes slow reorientations at timescales comparable to folded proteins. The slow motions are not perturbed by mutations in alpha-synuclein, which are related to genetic forms of Parkinson's disease, and do not depend on secondary and tertiary structural propensities. Ensemble-based hydrodynamic calculations suggest that the timescale of the underlying correlated motion is largely determined by hydrodynamic coupling between locally rigid segments. Our study indicates that long-range correlated dynamics are an intrinsic property of IDPs and offers a general physical mechanism of correlated motions in highly flexible biomolecular systems.
    Journal of the American Chemical Society 10/2014; · 10.68 Impact Factor
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    ABSTRACT: Bioinorganic chemistry of alpha-synuclein: Cu(I) binds preferentially to the N-terminal region, where metal-protein interactions at the Met motif and His-50 sites might explain the link between oxidative damage and protein aggregation.
    Journal of Inorganic Biochemistry 09/2014; · 3.20 Impact Factor
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    ABSTRACT: In a conformational selection scenario, manipulating the populations of binding-competent states should be expected to affect protein binding. We demonstrate how in silico designed point mutations within the core of ubiquitin, remote from the binding interface, change the binding specificity by shifting the conformational equilibrium of the ground-state ensemble between open and closed substates that have a similar population in the wild-type protein. Binding affinities determined by NMR titration experiments agree with the predictions, thereby showing that, indeed, a shift in the conformational equilibrium enables us to alter ubiquitin’s binding specificity and hence its function. Thus, we present a novel route towards designing specific binding by a conformational shift through exploiting the fact that conformational selection depends on the concentration of binding-competent substates.
    Angewandte Chemie 08/2014;
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    ABSTRACT: Residual dipolar couplings (RDCs) have proven to be an invaluable anisotropic NMR parameter for the structural elucidation of complex biopolymers and organic molecules. However, a remaining bottleneck limiting its wider use by organic and natural product chemists is the lack of a range of easily applicable aligning media for diverse organic solvents. In this study, graphene oxide (GO) liquid crystals (LCs) were developed to induce partial orientation of organic molecules allowing RDC measurements. These LCs were determined to be maintainable at very low concentrations (as low as 1 mg/mL, corresponding to a quadrupolar 2H splitting ranging from 2.8 Hz to 30 Hz and 13C-1H dipolar couplings of a maximum of 20 Hz for camphor in CH3COCH3/water system) and to be remarkably stable and broadly compatible with aqueous and organic solvents such as DMSO, CH3COCH3 and CH3CN. Moreover, compared to other alignment media, very clean and high-quality NMR spectra were acquired with the GO molecules in solution because of their rigidity and large molecular weight. The developed medium offers a versatile and robust method for RDC measurements that may routinize the RDC-based structure determination of organic molecules.
    Journal of the American Chemical Society 07/2014; · 10.68 Impact Factor
  • Alzheimer's and Dementia 07/2014; 10(4):P236-P237. · 17.47 Impact Factor
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    ABSTRACT: Alpha-Synuclein (αS) misfolding is associated with Parkinson's disease (PD) but little is known about the mechanisms underlying αS toxicity. Increasing evidence suggests that defects in membrane transport play an important role in neuronal dysfunction. Here we demonstrate that the GTPase Rab8a interacts with αS in rodent brain. NMR spectroscopy reveals that the C-terminus of αS binds to the functionally important switch region as well as the C-terminal tail of Rab8a. In line with a direct Rab8a/αS interaction, Rab8a enhanced αS aggregation and reduced αS-induced cellular toxicity. In addition, Rab8 - the Drosophila ortholog of Rab8a - ameliorated αS-oligomer specific locomotor impairment and neuron loss in fruit flies. In support of the pathogenic relevance of the αS-Rab8a interaction, phosphorylation of αS at S129 enhanced binding to Rab8a, increased formation of insoluble αS aggregates and reduced cellular toxicity. Our study provides novel mechanistic insights into the interplay of the GTPase Rab8a and αS cytotoxicity, and underscores the therapeutic potential of targeting this interaction.
    Neurobiology of Disease 06/2014; · 5.62 Impact Factor
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    ABSTRACT: We applied for the first time an innovative ligand-based NMR methodology (STI) to a medicinal-chemistry project aimed at the development of inhibitors for the enzyme 1-deoxy-D-xylulose-5-phosphate synthase (DXS). DXS is the first enzyme of the 2C-methyl-D-erythritol-4-phosphate (MEP) pathway, present in most bacteria (and not in humans) and responsible for the synthesis of the essential isoprenoid precursors. We designed de novo a first generation of fragments, using Deinococcus radiodurans DXS as a model enzyme, targeting the thiamine diphosphate (TDP) pocket of DXS whilst also exploring the putative substrate-binding pocket, where selectivity over other human TDP-dependent enzymes could be gained. The STI methodology – suitable for weak binders – was essential to determine the binding mode in solution of one of the fragments, circumventing the requirement for an X-ray co-crystal structure, which is known to be particularly challenging for this specific enzyme and in general for weak binders. Based on this finding, we carried out fragment growing and optimisation, which led to a three-fold more potent fragment, about as potent as the well-established thiamine analogue deazathiamine. The STI methodology proved therefore its strong potential as a tool to support medicinal-chemistry projects in their early stages, especially when dealing with weak binders.
    Chemical Science 06/2014; · 8.31 Impact Factor
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    ABSTRACT: Nuclear magnetic resonance (NMR) is a powerful tool for observing the motion of biomolecules at the atomic level. One technique, the analysis of relaxation dispersion phenomenon, is highly suited for studying the kinetics and thermodynamics of biological processes. Built on top of the relax computational environment for NMR dynamics is a new dispersion analysis designed to be comprehensive, accurate and easy-to-use. The software supports more models, both numeric and analytic, than current solutions. An automated protocol, available for scripting and driving the graphical user interface (GUI), is designed to simplify the analysis of dispersion data for NMR spectroscopists. Decreases in optimization time are granted by parallelization for running on computer clusters and by skipping an initial grid search by using parameters from one solution as the starting point for another -using analytic model results for the numeric models, taking advantage of model nesting, and using averaged non-clustered results for the clustered analysis. The software relax is written in Python with C modules and is released under the GPLv3+ license. Source code and precompiled binaries for all major operating systems are available from http://www.nmr-relax.com. edward@nmr-relax.com.
    Bioinformatics 04/2014; · 5.47 Impact Factor
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    ABSTRACT: Amyloid aggregation of α-synuclein (AS) has been linked to the pathological effects associated with Parkinson's disease (PD). Cu(II) binds specifically at the N-terminus of AS and triggers its aggregation. Site-specific Cu(I)-catalyzed oxidation of AS has been proposed as a plausible mechanism for metal-enhanced AS amyloid formation. In this study, Cu(I) binding to AS was probed by NMR spectroscopy, in combination with synthetic peptide models, site-directed mutagenesis, and C-terminal-truncated protein variants. Our results demonstrate that both Met residues in the motif (1)MDVFM(5) constitute key structural determinants for the high-affinity binding of Cu(I) to the N-terminal region of AS. The replacement of one Met residue by Ile causes a dramatic decrease in the binding affinity for Cu(I), whereas the removal of both Met residues results in a complete lack of binding. Moreover, these Met residues can be oxidized rapidly after air exposure of the AS-Cu(I) complex, whereas Met-116 and Met-127 in the C-terminal region remain unaffected. Met-1 displays higher susceptibility to oxidative damage compared to Met-5 because it is directly involved in both Cu(II) and Cu(I) coordination, resulting in closer exposure to the reactive oxygen species that may be generated by the redox cycling of copper. Our findings support a mechanism where the interaction of AS with copper ions leads to site-specific metal-catalyzed oxidation in the protein under physiologically relevant conditions. In light of recent biological findings, these results support a role for AS-copper interactions in neurodegeneration in PD.
    Inorganic Chemistry 04/2014; · 4.59 Impact Factor
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    ABSTRACT: The voltage-dependent anion channel (VDAC) mediates and gates the flux of metabolites and ions across the outer mitochondrial membrane (OMM) and is a key player in cellular metabolism and apoptosis. Here we characterized the binding of nucleotides to human VDAC1 (hVDAC1) on a single-residue level using NMR spectroscopy and site-directed mutagenesis. We find that hVDAC1 possesses one major binding region for ATP, UTP and GTP, which partially overlaps with a previously determined NADH-binding site. This nucleotide binding region is formed by the N-terminal α-helix, the linker connecting the helix to the first β-strand and adjacent barrel residues. hVDAC1 preferentially binds the charged forms of ATP, providing support for a mechanism of metabolite transport, in which direct binding to the charged form exerts selectivity, while at the same time permeation of the Mg(2+)-complexed ATP form is possible.
    Journal of Biological Chemistry 03/2014; · 4.65 Impact Factor
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    Acta Neuropathologica 03/2014; · 9.73 Impact Factor
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    ABSTRACT: The quality of genetically encoded calcium indicators (GECIs) has improved dramatically in recent years, but high-performing ratiometric indicators are still rare. Here we describe a series of fluorescence resonance energy transfer (FRET)-based calcium biosensors with a reduced number of calcium binding sites per sensor. These 'Twitch' sensors are based on the C-terminal domain of Opsanus troponin C. Their FRET responses were optimized by a large-scale functional screen in bacterial colonies, refined by a secondary screen in rat hippocampal neuron cultures. We tested the in vivo performance of the most sensitive variants in the brain and lymph nodes of mice. The sensitivity of the Twitch sensors matched that of synthetic calcium dyes and allowed visualization of tonic action potential firing in neurons and high resolution functional tracking of T lymphocytes. Given their ratiometric readout, their brightness, large dynamic range and linear response properties, Twitch sensors represent versatile tools for neuroscience and immunology.
    Nature Methods 01/2014; · 23.57 Impact Factor
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    ABSTRACT: The traditional view of how intracellular effector proteins are recruited to the B cell antigen receptor (BCR) complex at the plasma membrane is based on the occurrence of direct protein-protein interactions, as exemplified by the recruitment of the tyrosine kinase Syk (spleen tyrosine kinase) to phosphorylated motifs in BCR signaling subunits. By contrast, the subcellular targeting of the cytosolic adaptor protein SLP-65 (Src homology 2 domain-containing leukocyte adaptor protein of 65 kD), which serves as a proximal Syk substrate, is unclear. We showed that SLP-65 activation required its association at vesicular compartments in resting B cells. A module of ~50 amino acid residues located at the amino terminus of SLP-65 anchored SLP-65 to the vesicles. Nuclear magnetic resonance spectroscopy showed that the SLP-65 amino terminus was structurally disordered in solution but could bind in a structured manner to noncharged lipid components of cellular membranes. Our finding that preformed vesicular signaling scaffolds are required for B cell activation indicates that vesicles may deliver preassembled signaling cargo to sites of BCR activation.
    Science Signaling 01/2014; 7(339):ra79. · 7.65 Impact Factor
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    ABSTRACT: Here we report the synthesis of all four stereoisomers of mefloquine. Mefloquine (Lariam) is an important anti-malaria drug that is applied as a racemate of the erythro form. However, the (-)-isomer induces psychosis, while the (+)-enantiomer does not have this undesired side effect. There are six syntheses of which five lead to the wrong enantiomer without the authors of these syntheses noting that they had synthesized the wrong compound. At the same time physical chemistry investigations had assigned the absolute configuration correctly and the last enantioselective synthesis that took these results into account delivered the correct absolute configuration. Since various synthetic approaches failed to provide the correct stereoisomers in previous syntheses, we submit here a synthetic approach with a domino Sonogashira-6π-electrocyclisation as key step that confirmed synthetically the correct absolute configuration of all four isomers.
    Chemistry - A European Journal 11/2013; · 5.93 Impact Factor
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    ABSTRACT: Ag-mediated B cell stimulation relies on phospholipase Cγ2 (PLCγ2) for Ca(2+) mobilization. Enzymatic activity of PLCγ2 is triggered upon Src homology 2 domain-mediated binding to the tyrosine-phosphorylated adaptor SLP65. However, SLP65 phosphorylation outlasts the elevation of cytosolic Ca(2+) concentration suggesting additional levels of PLCγ2 regulation. We show in this article that the functionality of the PLCγ2/SLP65 complex is controlled by the weakly characterized C2 domain of PLCγ2. Usually C2 domains bind membrane lipids, but that of PLCγ2 docks in a Ca(2+)-regulated manner to a distinct phosphotyrosine of SLP65. Hence, early Ca(2+) fluxing provides feed-forward signal amplification by promoting anchoring of the PLCγ2 C2 domain to phospho-SLP65. As the cellular Ca(2+) resources become exhausted, the concomitant decline of Ca(2+) dampens the C2-phosphotyrosine interaction so that PLCγ2 activation terminates despite sustained SLP65 phosphorylation.
    The Journal of Immunology 10/2013; · 5.52 Impact Factor
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    ABSTRACT: An ensemble-based approach is presented to explore the conformational space sampled by a multi-domain protein showing moderate inter-domain dynamics in terms of translational and rotational motions. The strategy was applied on a complex of calmodulin (CaM) with the IQ-recognition motif from the voltage-gated calcium channel Cav1.2 (IQ), which adopts three different inter-domain orientations in the crystal. The N60D mutant of calmodulin was used to collect pseudocontact shifts (pcs's) and paramagnetically induced residual dipolar couplings (RDC's) for six different lanthanide ions. Then, starting from the crystal structure, pools of conformations were generated by free MD. We found the three crystal conformations in solution, but four additional MD derived conformations had to be included into the ensemble to fulfill all the paramagnetic data and cross validate optimally against unused paramagnetic data. Alternative approaches lead to similar ensembles. Our "ensemble" approach is a simple and efficient tool to probe and describe the inter-domain dynamics and represents a general method that can be used to provide a proper ensemble description of multi-domain proteins.
    Journal of the American Chemical Society 10/2013; · 10.68 Impact Factor
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    ABSTRACT: The cytoplasmic PASC domain of the fumarate responsive sensor kinase DcuS of Escherichia coli links the transmembrane to the kinase domain. PASC is also required for interaction with the transporter DctA serving as a cosensor of DcuS. Earlier studies suggested that PASC functions as a hinge and transmits the signal to the kinase. Reorganizing the PASC dimer interaction and, independently, removal of DctA, converts DcuS to the constitutive ON state (active without fumarate stimulation). ON mutants were categorized with respect to these two biophysical interactions and the functional state of DcuS: type I-ON mutations grossly reorganize the homodimer, and decrease interaction with DctA. Type IIA-ON mutations create the ON state without grossly reorganizing the homodimer, whereas interaction with DctA is decreased. The type IIB-ON mutations were neither in PASC /PASC , nor in DctA/DcuS interaction affected, similar to fumarate activated wild-typic DcuS. OFF mutations never affected dimer stability. The ON mutations provide novel mechanistic insight: PASC dimerization is essential to silence the kinase. Reorganizing the homodimer and its interaction with DctA activate the kinase. The study suggests a novel ON homo-dimer conformation (type IIB) and an OFF conformation for PASC . Type IIB-ON corresponds to the fumarate induced wild-type conformation, representing an interesting target for structural biology.
    MicrobiologyOpen. 09/2013;

Publication Stats

7k Citations
1,728.97 Total Impact Points

Institutions

  • 2000–2014
    • Max Planck Institute for Biophysical Chemistry
      • Department of NMR-based Structural Biology
      Göttingen, Lower Saxony, Germany
  • 2013
    • Mahidol University
      • Department of Chemistry
      Bangkok, Bangkok, Thailand
  • 2005–2013
    • Johannes Gutenberg-Universität Mainz
      • • Institute of Microbiology and Wine Research
      • • Institute of Organic Chemistry
      Mainz, Rhineland-Palatinate, Germany
  • 2012
    • Ludwig-Maximilians-University of Munich
      München, Bavaria, Germany
    • Bar Ilan University
      Gan, Tel Aviv, Israel
    • The University of Edinburgh
      Edinburgh, Scotland, United Kingdom
  • 2006–2012
    • Rosario National University
      • Instituto de Biología Molecular y Celular de Rosario (IBR)
      Rosario, Provincia de Santa Fe, Argentina
    • Imperial College London
      • Department of Chemistry
      London, ENG, United Kingdom
  • 1992–2012
    • Goethe-Universität Frankfurt am Main
      • • Institute of Biochemistry
      • • Institut für Organische Chemie und Chemische Biologie
      Frankfurt, Hesse, Germany
  • 2011
    • Florida State University
      • Department of Chemistry and Biochemistry
      Tallahassee, FL, United States
  • 2010
    • Paul-Ehrlich-Institut
      Langen, Hesse, Germany
  • 2001–2010
    • Massachusetts Institute of Technology
      • Department of Chemistry
      Cambridge, Massachusetts, United States
  • 2009
    • Universität Basel
      Bâle, Basel-City, Switzerland
    • University of Santiago de Compostela
      • Facultad de Química
      Santiago de Compostela, Galicia, Spain
  • 2008
    • Max Planck Research Unit for Structural Molecular Biology at DESY
      Hamburg, Hamburg, Germany
    • University of Alberta
      • Department of Biochemistry
      Edmonton, Alberta, Canada
    • Texas A&M University
      • Department of Chemistry
      College Station, TX, United States
    • Georg-August-Universität Göttingen
      Göttingen, Lower Saxony, Germany
  • 2007
    • Max Planck Institute for Developmental Biology
      Tübingen, Baden-Württemberg, Germany
  • 2000–2005
    • Philipps-Universität Marburg
      • • Fachbereich Chemie
      • • Faculty of Biology
      Marburg an der Lahn, Hesse, Germany
  • 2004
    • Max Planck Institute for Terrestrial Microbiology
      Marburg, Hesse, Germany
  • 2001–2004
    • Technische Universität München
      • Abteilung Organische Chemie und Biochemie
      München, Bavaria, Germany
  • 1994–2004
    • University Hospital Frankfurt
      Frankfurt, Hesse, Germany
    • University of Oxford
      Oxford, England, United Kingdom
  • 2003
    • Clark University
      Worcester, Massachusetts, United States
  • 2002
    • Humboldt-Universität zu Berlin
      • Department of Chemistry
      Berlin, Land Berlin, Germany
  • 1986–1993
    • ETH Zurich
      • Laboratory of Physical Chemistry
      Zürich, ZH, Switzerland
  • 1987–1991
    • Hochschule für Technik Zürich
      Zürich, Zurich, Switzerland