Whole proteome analyses on Ruminiclostridium cellulolyticum show a modulation of the cellulolysis machinery in response to cellulosic materials with subtle differences in chemical and structural properties

Lignocellulosic materials from municipal solid waste emerge as attractive resources for anaerobic digestion biorefinery. To increase the knowledge required for establishing efficient bioprocesses, dynamics of batch fermentation by the cellulolytic bacterium Ruminiclostridium cellulolyticum were compared using three cellulosic materials, paper handkerchief, cotton discs and Whatman filter paper. Fermentation of paper handkerchief occurred the fastest and resulted in a specific metabolic profile: it resulted in the lowest acetate-to-lactate and acetate-to-ethanol ratios. By shotgun proteomic analyses of paper handkerchief and Whatman paper incubations, 151 proteins with significantly different levels were detected, including 20 of the 65 cellulosomal components, 8 non-cellulosomal CAZymes and 44 distinct extracytoplasmic proteins. Consistent with the specific metabolic profile observed, many enzymes from the central carbon catabolic pathways had higher levels in paper handkerchief incubations. Among the quantified CAZymes and cellulosomal components, 10 endoglucanases mainly from the GH9 families and 7 other cellulosomal subunits had lower levels in paper handkerchief incubations. An in-depth characterization of the materials used showed that the lower levels of endoglucanases in paper handkerchief incubations could hypothetically result from its lower crystallinity index (50%) and degree of polymerization (970). By contrast, the higher hemicellulose rate in paper handkerchief (13.87%) did not result in the enhanced expression of enzyme with xylanase as primary activity, including enzymes from the xyl-doc cluster. It suggests the absence, in this material, of molecular structures that specifically lead to xylanase induction. The integrated approach developed in this work shows that subtle differences among cellulosic materials regarding chemical and structural characteristics have significant effects on expressed bacterial functions, in particular the cellulolysis machinery, resulting in different metabolic patterns and degradation dynamics.This work was supported by a grant [R2DS 2010-08] from Conseil Regional d'Ile-de-France through DIM R2DS programs (http://www.r2ds-ile-de-france.com/). Irstea (www.irstea.fr/) contributed to the funding of a PhD grant for the first author. The funders provided support in the form of salaries for author [NB], funding for consumables and laboratory equipment, but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. Omics Services provided support in the form of salaries for authors [VS, MD], but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors [NB, VS, MD] are articulated in the 'author contributions' section.info:eu-repo/semantics/publishedVersio

Tomographie de fluorescence DUV sur des systèmes d’intérêt biologique

L’utilisation de la lumière synchrotron en ultraviolet lointain (DUV, de longueur d’onde comprise entre 120 et 300 nm) pour exciter la fluorescence des molécules en microscopie, permet de s’affranchir de marqueurs de contrastes. Un microscope inversé DUV est installé à SOLEIL sur la ligne de lumière DISCO. La motorisation axiale de l’échantillon et le traitement du signal par déconvolution rendent possible l’obtention de tomogrammes avec des résolutions de l’ordre de 100 nm. Deux applications biologiques sont présentées ici : l’activité d’une enzyme, l’amylase, sur un grain d’amidon; et le suivi de l’entrée de nanoparticules dans des cellules cancéreuses

Chemo-enzymatic synthesis of xylogluco-oligosaccharides and their interactions with cellulose

International audienceA novel class of complex oligosaccharides with a backbone made of twelve beta-(1 -> 4)-D-glucosyl units that may be regularly substituted on their primary position with alpha-D-xylosyl or beta-D-(1 -> 2)-galactosyl-alpha-D-xylosyl residues were readily synthesized using enzymatic hydrolysis of the plant polysaccharide xyloglucan (XG), followed by chemical modifications of the well-defined oligosaccharides and their enzymatic coupling catalyzed by the Cel7B E197A glycosynthase from Humicola insolens. These complex oligosaccharides proved useful for a better understanding of the effects of structural variation of xyloglucan on interactions with bacterial microcrystalline cellulose (BMCC)