Plant cell wall material is the most abundant source of organic carbon in the biosphere, and its enzymatic degradation represents an important source of nutrients for microorganisms, plants, and herbivores. Plant cell wall-degrading enzymes are widely used in the biotechnology sector for the production of detergents, paper, textiles, and animal and human foods, as well as for the production of renewable biofuels. Therefore, the discovery of new and more efficient plant cell wall-degrading enzymes can potentially have numerous and important biotechnology applications.
Because of its complex chemical structure, enzymatic attack of the plant cell wall presents a significant challenge, and microorganisms usually rely on an extensive repertoire of enzymes to efficiently degrade complex plant cell wall materials. In order to obtain a complete picture of the plant cell wall-degrading apparatus, we completely sequenced, annotated, and analyzed the genome of one of the most efficient plant cell wall degraders, C. japonicus. The results presented here show that the genome of C. japonicus is remarkably similar to that of the gram-negative marine bacterium S. degradans 2-40T. Both C. japonicus and S. degradans produce a repertoire of glycoside hydrolases, lyases, and esterases that can degrade cellulose, xylan, mannan, chitin, pectin, starch, and laminarin, with GH13 and GH43 being two of the largest enzyme families.
The large number of GH43 glycoside hydrolases is an intriguing characteristic of C. japonicus
= 14), a feature shared with the colonic bacterium Bacteroides thetaiotaomicron
= 33) (44
), as well as S. degradans
= 13) and some aerobic fungi, including Aspergillus. B. thetaiotaomicron
utilizes complex carbohydrates as major nutrients. However, rather than attacking the internal regions of integral plant structural polysaccharides (such as cellulose and hemicellulose), the vast majority of glycoside hydrolases are predicted to be exo-acting, e.g., removing monosaccharides from the complex glycans presented on the surface of mammalian cells and the easily accessible plant polysaccharides such as the pectins. Thus, these extensive, and highly accessible, pectin decorations are likely to represent an important source of nutrients for both colonic bacteria and soil saprophytic microorganisms that use plant biomass as an important nutrient source.
The complement of cellulases encoded by the genome of C. japonicus is similar to that of S. degradans, which also contains a single cellobiohydrolase from GH6. However, both bacteria lack a reducing-end cellobiohydrolase, which is considered an important component of fungal (GH7) and clostridial (GH48) cellulose degradative systems. As both S. degradans and C. japonicus efficiently degrade crystalline cellulose, it would appear that not all enzyme systems which hydrolyze this important polysaccharide require a GH7 or GH48 exo-acting glycoside hydrolase.
Examination of the C. japonicus
CBM repertoire shows a dominance of CBM2 and CBM10 modules. This observation likely reflects specificity for crystalline cellulose, which, because of its prevalence in all plant cell walls, functions as a universal receptor for these degradative enzymes, as suggested previously (18
). As described above, the plant cell wall hydrolases of C. japonicus
contain CBMs from numerous families. In sharp contrast, the corresponding fungal enzymes generally contain only a family 1 CBM, suggesting that targeting these glycoside hydrolases to polysaccharides other than crystalline cellulose does not confer an obvious evolutionary advantage. In the case of C. japonicus
, it is apparent that the different CBMs appended to the catalytic modules of the glycoside hydrolases will target these enzymes to diverse plant cell wall components, providing an explanation for the complex arrangement of these modules in the degradative enzymes.
Approximately 50% of the C. japonicus
plant-degradative apparatus appears to be shared with S. degradans
, consistent with the observation that the cell walls of terrestrial and marine plants have many polysaccharides in common. Although these two species share this high level of similarity in their substrate utilization machinery, it is still not clear whether the common ancestor of C. japonicus
and S. degradans
was a marine or a terrestrial bacterium, and additional analyses of their genome sequences might reveal what adaptations underlie these different lifestyles. For example, consistent with the marine environment of S. degradans
, the bacterium produces an extensive array of agarases and carageenases, while the terrestrial C. japonicus
lacks these enzymes. Agarases are found mostly in marine habitats. This is consistent with the fact that agar, being a product of marine algae, is available to and utilized by marine organisms, such as S. degradans
, as a convenient carbon and energy source. Mannan, however, is likely to be more common in terrestrial plants than in salt marsh cord grass (8
), and consistent with this, C. japonicus
has a more extended mannan-degrading system than S. degradans
. It is also apparent that while both organisms contain a large number of CBMs, there has been an extensive expansion of CBM6 and CBM32 modules in S. degradans
, which likely reflects the diversity of sugars present in the marine-specific polysaccharides. These differences highlight how the selection pressures imposed by terrestrial and marine environments have influenced the evolution of the plant cell wall-degrading apparatus of saprophytic prokaryotes.