Dietary fibres of plant cell walls are degraded and fermented by fibrolytic bacteria in the human gut. Xylan being one of the major polysaccharides constituting the plant cell walls, we have characterised two bacteria presenting important xylanolytic activities: Bacteroides xylanisolvens XB1AT and Roseburia intestinalis XB6B4. The xylanolytic activities of these species grown on various substrates rich in xylan were analysed qualitatively and quantitatively. The two bacteria degraded and fermented all the substrates tested very efficiently, but they colonised them differently. The enzymatic system of these strains, studied by zymogram, revealed a large number of xylanases of molecular mass comprised between 37 and 170kDa. Specific activities of protein extracts were very high, and were mainly associated with the bacteria. The xylanase-encoding genes of the two bacteria studied, and of another xylanolytic bacterium, Bacteroides sp. XB12B, were identified. They encode xylanases from glycoside-hydrolase families 10 and 43, and are modular. Some xylanases were also identified by proteomics. The conditions for expression of some of the genes were studied by quantitative PCR, and showed a regulation by xylan. Several clusters of genes predicted to code for hemicellulases were found in the genome of B. xylanisolvens XB1AT. Finally, a 38kDa GH10 xylanase produced by B. xylanisolvens XB1AT, named Xyn10A, was characterized. The xyn10A gene was cloned and expressed in E. coli. The recombinant Xyn10A was very active against xylan and xylooligosaccharides, and also showed a weak CMC activity. Xyn10A may be periplasmic in B. xylanisolvens XB1AT. This work represents a major contribution to our knowledge of the mechanisms of xylan degradation by representatives of the Bacteroides and Roseburia genera, and also a progress in assessing their respective roles in the human colon ecosystem.
