Solid-Phase Cross-Linking (SPCL): A new tool for protein structure studies

International audienceA wide range of chemical reagents are available to study the protein-protein interactions or protein structures. After reaction with such chemicals, covalently modified proteins are digested, resulting in shorter peptides that are analyzed by mass spectrometry (MS). Used especially when NMR of X-ray data are lacking, this methodology requires the identification of modified species carrying relevant information, among the unmodified peptides. To overcome the drawbacks of existing methods, we propose a more direct strategy relying on the synthesis of solid-supported cleavable monofunctional reagents and cross-linkers that react with proteins and that selectively release, after protein digestion and washings, the modified peptide fragments ready for MS analysis. Using this Solid-Phase Cross-Linking (SPCL) strategy, only modified sequences are analyzed and consistent data can be easily obtained since the signals of interest are not masked or suppressed by over-represented unmodified materials

A new generation of cross-linkers for selective detection by MALDI MS

International audienceWe designed a new cross-linker bearing a CHCA moiety. The use of the CHCA-tagged crosslinker JMV 3378 in conjunction with a neutral MALDI matrix a-cyano-4-hydroxycinnamic methyl ester enabled specific signal enhancement in MALDI-TOF MS of cross-link containing peptides. Discrimination between modified and non-modified peptides can be achieved by comparison of two spectra, one using CHCA and the other using the a-cyano-4- hydroxycinnamic methyl ester matrix. The methodology was validated using cytochrome c and apo-myoglobine as model proteins

Enhanced structural and functional genome elucidation of the arsenite-oxidizing strain Herminiimonas arsenicoxydans by proteomics data.

International audienceThe arsenite-oxidizing strain Herminiimonas arsenicoxydans proteome was investigated with gel electrophoresis and tandem mass spectrometry analyses. The comparison of experimental and theoretical M(r) and pI, as well as that of peptide sequences identified by MS and predicted protein sequences, allowed the correction of five protein annotations. More importantly, the functional analysis of SDS- and 2D-PAGE proteome maps obtained in the presence of arsenic, combined with partial transcriptomic results indicate that H. arsenicoxydans expressed genes and proteins required not only for arsenic detoxification or stress response but also involved in motility, exopolysaccharide synthesis, phosphate import or energetic metabolism. This study provides therefore new insights into the adaptation processes of H. arsenicoxydans in response to arsenic

Analysis of XFEL serial diffraction data from individual crystalline fibrils

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