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

Valorization, comparison and characterization of coconuts waste and cactus in a biorefinery context using NaClO2-C2H4O2 and sequential NaClO2-C2H4O2/autohydrolysis pretreatment

The search for new sources of lignocellulosic raw materials for the generation of energy and new compounds encourages the search for locations not well known and with a high potential for biomass availability as is the case of the Northeast Region of Brazil. Thus, the cactus (CAC), green coconut shell (GCS), mature coconut fibre and mature coconut shell were pretreated by NaClO2C2H4O2 and sequential NaClO2C2H4O2/autohydrolysis aiming at the obtention of high added-value compounds in the liquid fraction and solid phase. The yield of the solid phase was between 61.42 and 90.97% and the reduction up to 91.63% of lignin in the materials pretreated by NaClO2C2H4O2. After NaClO2C2H4O2/autohydrolysis pretreatment the obtained solids yield was between 43.57 and 52.08%, with a solubilization of the hemicellulose content up to 81.42%. For both pretreatments the cellulosic content remained almost unchanged. The pretreated solids were characterized by SEM, X-ray and crystallinity indexes showing significant modifications when submitted to pretreatments. These results were further confirmed by the enzymatic conversion yields of 81.6890.03 and 86.9790.36% of the LCMs pretreated by NaClO2C2H4O2 and pretreated by NaClO2C2H4O2/autohydrolysis, respectively. The resulting liquors had a total phenolic compounds content between 0.20 and 3.05 g/L, lignin recovered up to 7.40 g/L (absence of sulphur) and xylooligosaccharides between 16.13 and 20.37 g/L. Thus, these pretreatments showed an efficient fractionation of LCMs, especially in the GCS, being an important requirement for the generation of products and byproducts in the context of the biorefinery.The authors gratefully acknowledge the Brazilian research funding agencies CNPq and CAPES for financial support. Financial support from the Energy Sustainability Fund 2014-05 (CONACYT-SENER), Mexican Centre for Innovation in Bioenergy (CemieBio), Cluster of Bioalcohols (Ref. 249564) is gratefully acknowledged. We also gratefully acknowledge support for this research by the Mexican Science and Technology Council (CONACYT, Mexico) for the infrastructure project - INFR201601 (Ref. 269461) and CB-2015-01 (Ref. 254808).info:eu-repo/semantics/publishedVersio