1-06: Bacterial community dynamics and glycoside hydrolase activities during thermophilic biomass deconstruction

Bacteria from thermophilic environments are an emerging source of enzymes for deconstruction of biomass to produce biofuels. Identifying specific bacterial groups that produce enzymes and understanding the mechanisms through which these groups deconstruct biomass is critical to develop mixtures of enzymes that may be applied to the industrial production of biofuels. To identify these enzymes, we have focused on adapting thermophilic bacterial microbial communities to specific biomass feedstocks and pretreatment conditions, followed by microbial community analysis and functional characterization of glycoside hydrolases. Multiple enrichments have demonstrated that populations related to gram-positive Firmicutes (Thermobacillus, Paenibacillus, Brevibacillus), thermophilic Bacteriodetes (Rhodothermus, Chitinophaga) and Thermus are ubiquitous in these enrichments. Using community proteogenomics and targeted isolations, we have demonstrated that the Firmicutes are responsible for initial deconstruction of insoluble substrates and the Bacteriodetes and Thermus populations consume the soluble hydrolysis products of initial polysaccharide deconstruction. Assessment of individual thermophilic gram-positive isolates has demonstrated that these bacteria possess diverse mechanisms for hydrolyzing polysaccharides, including secretion of individual proteins and the display of multi-enzyme complexes on bacterial surfaces. Knowledge of the bacterial polysaccharide hydrolysis mechanisms has enabled the design of thermophilic enzymatic cocktails for biomass deconstruction. These cocktails are a combination of supernatants from the enrichments and recombinant enzymes obtained by heterologous expression of glycoside hydrolases from metagenomic sequencing. This work has provided thermophilic enzymatic cocktails that efficiently hydrolyze ionic-liquid pretreated switchgrass at 80 °C.