10-01: Screening for novel biomass-degrading enzymes in environmental samples

In natural ecosystems, microbial communities may share the task of breaking down lignocellulosic biomass by producing degradative enzymes that act in concert. Given that a majority of microbial species are still undescribed, we can hypothesize that many of their enzymes remain unknown. The production of cellulosic ethanol depends on the efficient breakdown of biomass feedstocks, and the need for new enzymes for this capacity is pertinent. As part of the Bioenergy Science Center we are exploring environmental, cellulolytic, microbial communities to discover novel glycosyl hydrolases. The samples we investigated include microbial communities from herbivorous animal rumen and caecum, wood-eating beetle larvae guts, and Biotraps, in situ microbial enrichments, from temperate and high-temperature environments. By accessing the metagenome of each sample, using culture-independent screening methods, we can discover the genes and enzymes from all the microbes within each community. Each sample community was analyzed, using ribosomal markers, for microbial composition and abundance. The data obtained help to inform decisions on choosing the samples to screen, the most beneficial screening methods, and the depth of library screening coverage. To discover the broadest array of enzymes, we utilized two distinct, complementary screening methods. First, DNA libraries, created from sample metagenomic DNA, were screened for active enzymes using Verenium's proprietary, high-throughput, Gigamatrix® platform. The second, sequence-based method, utilized degenerate PCR primers to screen sample metagenomic DNA for family 48 cellobiohydrolases. We combined Verenium's knowledge and expertise in community microbial exploration, sample bacterial composition analyses, and high-throughput discovery methods, to further the advancement of cellulosic ethanol technology.