A5028010948- Protein Structure and Dynamics
- Enzyme Structure and Function
- Photosynthetic Processes and Mechanisms
- Bacterial Genetics and Biotechnology
- Peptidase Inhibition and Analysis
- Glycosylation and Glycoproteins Research
- RNA and protein synthesis mechanisms
- Advanced Fluorescence Microscopy Techniques
- Bacteriophages and microbial interactions
- Cancer Research and Treatments
- Photoreceptor and optogenetics research
- Pneumonia and Respiratory Infections
- Molecular spectroscopy and chirality
- Hemoglobin structure and function
- bioluminescence and chemiluminescence research
- RNA Research and Splicing
- Nuclear Structure and Function
- HIV Research and Treatment
- Carbohydrate Chemistry and Synthesis
- Streptococcal Infections and Treatments
- Porphyrin Metabolism and Disorders
- Advanced Electron Microscopy Techniques and Applications
- Metabolomics and Mass Spectrometry Studies
- Bacterial Infections and Vaccines
- Electron and X-Ray Spectroscopy Techniques
Université Grenoble Alpes
2010-2025
Centre National de la Recherche Scientifique
2010-2025
Institut de Biologie Structurale
2013-2025
CEA Grenoble
2010-2025
Commissariat à l'Énergie Atomique et aux Énergies Alternatives
2010-2025
Université Joseph Fourier
2000-2013
Laboratoire des Biomolécules
2003-2004
Cyan variants of green fluorescent protein are widely used as donors in Förster resonance energy transfer experiments. The popular, but modestly bright, Enhanced Fluorescent Protein (ECFP) was sequentially improved into the brighter Super 3A (SCFP3A) and mTurquoise, latter exhibiting a high-fluorescence quantum yield long mono-exponential fluorescence lifetime. Here we combine X-ray crystallography excited-state calculations to rationalize these stepwise improvements. enhancement originates...
Sometimes less is more: [13C1H3]methyl isotopomers can be biosynthetically incorporated specifically into the pro-S methyl groups of leucine and valine residues in large protein assemblies within a perdeuterated background by using an acetolactate precursor. This stereospecific labeling strategy considerably enhances NMR spectra for assemblies. Detailed facts importance to specialist readers are published as ”Supporting Information”. Such documents peer-reviewed, but not copy-edited or...
Enhanced cyan fluorescent protein (ECFP) and its variant Cerulean are genetically encoded fluorophores widely used as donors in FRET-based cell imaging experiments. First, we have confirmed through denaturation experiments that the double-peak spectroscopic signature of these proteins originates from indole ring chromophore. Then, to explain improvement fluorescence properties compared those ECFP, determined high-resolution crystal structures two at physiological pH performed molecular...
Summary Bacterial division requires the co‐ordination of membrane invagination, driven by constriction FtsZ‐ring, and concomitant cell wall synthesis, performed high‐molecular‐weight penicillin‐binding proteins (HMW PBPs). Using immunofluorescence techniques, we show in Streptococcus pneumoniae that this PBP3, a d , ‐carboxypeptidase degrades substrate HMW PBPs. In mutant deprived apparent rings PBPs FtsZ are no longer co‐localized. wild‐type cells, PBP3 is absent at future site present over...
The green fluorescent protein (GFP) from the jellyfish Aequoria victoria has been shown to dimerize at high concentrations, which could lead artefacts in imaging experiments. To ensure a truly monomeric state, an A206K mutation introduced into most of its widely used variants, with minimal effect on spectroscopic properties. Here, first structure one these cyan mTurquoise, is presented and compared that dimeric version mTurquoise-K206A. No significant structural change detected chromophore...
Abstract The fungal Bromodomain and Extra‐Terminal (BET) protein Bdf1 is a potential antifungal target against invasive infections. However, the need to selectively inhibit both bromodomains (BDs) over human orthologs lack of molecular tools assess on‐target efficacy hamper efforts develop BD inhibitors as therapeutics. This study reports phenyltriazine compound that inhibits BDs from pathogen Candida glabrata with selectivity orthologous BET Brd4. On‐target activity established by devising...
Summary DivIB, DivIC and FtsL are bacterial proteins essential for cell division, which show interdependencies their stabilities localization. We have reconstituted in vitro a trimeric complex consisting of the recombinant extracellular domains three from Streptococcus pneumoniae. The domain DivIB was found to associate with heterodimer those FtsL. heterodimerization artificially constrained by fusion interacting coiled‐coils. Immunofluorescence experiments showed that is always localized at...
Abstract Background Streptococcus pneumoniae is a widely distributed commensal Gram-positive bacteria of the upper respiratory tract. Pneumococcal colonization can progress to invasive disease, and thus become lethal, reason why antibiotics vaccines are designed limit dramatic effects in such cases. As consequence, pneumococcus has developed efficient antibiotic resistance, use covering limited number serotypes as Pneumovax ® Prevnar results expansion non-covered serotypes. surface proteins...
Abstract Recent technical advances have revolutionized the field of cryo-electron microscopy (cryo-EM). However, most monomeric proteins remain too small (<100 kDa) for cryo-EM analysis. To overcome this limitation, we explored a strategy whereby target protein is genetically fused to homo-oligomeric scaffold and junction optimized allow adopt symmetry, thereby generating chimeric particle suitable cryo-EM. demonstrate concept, maltose-binding (MBP), 40 kDa monomer, glutamine synthetase,...
Synchrotrons are now producing thousands of macromolecular structures each year. The need for complementary techniques available on site has progressively emerged, either to assess the relevance structure a protein or monitor changes that may occur during X-ray diffraction data collection. Microspectrophotometers in UV–visible absorbance fluorescence mode have evolved over past few decades become instruments choice perform such tests. Described here recent improvements microspectrophotometer...
Abstract miniSOG, developed as the first fully genetically encoded singlet oxygen photosensitiser, has found various applications in cell imaging and functional studies. Yet, miniSOG suboptimal properties, including a low yield of generation, which can nevertheless be improved tenfold upon blue light irradiation. In previous study, we showed that this improvement was due to photolysis chromophore, flavin mononucleotide (FMN), into lumichrome, with concomitant removal phosphoribityl tail,...
Manchmal ist weniger mehr: 13C1H3-Methylisotopomere können biosynthetisch mithilfe einer Acetolactatvorstufe spezifisch als pro-S-Methylgruppen von Leucin- und Valinresten in ansonsten perdeuterierte Proteine eingefügt werden. Durch diese Markierungsstrategie wird die Qualität der NMR-Spektren großer Proteinassoziationen deutlich verbessert.
The synthesis of peptidoglycan, the major component bacterial cell wall, is essential to survival, yet its mechanism remains poorly understood. In present work, we have isolated several membrane protein complexes consisting late division proteins Streptococcus pneumoniae: DivIB, DivIC, FtsL, PBP2x and FtsW, or subsets thereof. We co-expressed from S. pneumoniae in Escherichia coli. By combining two successive affinity chromatography steps, obtained with a very good purity. These are...
ECFP, the first usable cyan fluorescent protein (CFP), was obtained by adapting tyrosine-based chromophore environment in green to that of a tryptophan-based one. This first-generation CFP superseded popular Cerulean, CyPet, and SCFP3A were engineered rational random mutagenesis, yet latter CFPs still exhibit suboptimal properties pH sensitivity reversible photobleaching behavior. These flaws serendipitously corrected third-generation mTurquoise its successors without an obvious rationale....
Membrane proteins constitute about one third of encoded by all genomes, but only a small percentage have their structures deposited in the Protein Data Bank. One bottleneck pipeline from expression to structure determination is identification detergents that maintain protein soluble, stable, and active state. Here, we describe small‐scale automated procedure easily rapidly screen for solubilization purification membrane proteins, perform detergent exchange, or identify conditions preserving...
Tetrahedral (TET) aminopeptidases are large polypeptide destruction machines present in prokaryotes and eukaryotes. Here, the rules governing their assembly into hollow 12-subunit tetrahedrons addressed by using TET2 from Pyrococcus horikoshii (PhTET2) as a model. Point mutations allowed capture of stable, catalytically active precursor. Small angle x-ray scattering revealed that it is dimer whose architecture solution identical to determined crystallography within fully assembled TET...