| Fossil fuels are a nonrenewable resource. Their wide geological distribution and relatively simple acquisition have allowed massive increases in human population and associated energy expenditure over the last century. The current rate of consumption and the expectation of reduced fuel supplies predicate the need to develop new energy sources that may be merged with the current energy infrastructure. As an underutilized renewable resource, lignocellulosic biomass may, through microbial bioconversion, contribute to the environmentally benign production of alternative fuels and chemical feedstocks. A major target for this conversion is methylglucuronoxylan, the predominant structural polysaccharide in the hemicellulose fraction of hardwood and crop residues. The research described herein has focused on xylanolytic bacteria and their secreted endoxylanases that are involved in the depolymerization of the methylglucuronoxylan. In this work, endoxylanases of glycosyl hydrolase families 5 and 10 (GH 5 and GH 10 xylanases) have been characterized with emphasis on the native bacterial host and utilization of the hydrolysis limit products of methylglucuronoxylan. Recombinant constructs of the genes encoding these xylanases have made them available for definitive characterization and for expression in transgenic organisms. XynA1, a multimodular GH 10 xylanase anchored to the cell surface of Paenibacillus sp. strain JDR-2, generates aldouronates that are efficiently assimilated and metabolized by this organism. The proximity between XynA1 and the native Paenibacillus host, and the proficient utilization of the resulting hydrolysis products, identify a process of vectoral assimilation of methylglucuronoxylan-derived products. A GH 5 xylanase encoded by the ynfF gene in Bacillus subtilis 168 is directed for catalysis by methylglucuronosyl substitutions on the xylan chain, supporting its application in an accessory role in the overall depolymerization process. Secretion of this GH 5 and a GH 11 endoxylanase by the genetically malleable B. subtilis 168, for which the entire genome has been sequenced, recommends it as a target for introduction of genes encoding the GH 10 endoxylanase, XynA1 and aldouronate-utilization enzymes for efficient depolymerization and metabolism of methylglucuronoxylan. These discoveries provide insight needed for the development of second-generation bacterial biocatalysts for the direct conversion of lignocellulosic biomass to alternative fuels and bio-based products. |
