P30: Biocatalysts for biomass degradation from the compost metagenome

Microorganisms from natural environments are a rich source of new biocatalysts and recent advances in DNA sequencing technology have made the recovery of large numbers of gene sequences from the environment feasible.  However, a major bottleneck still exists - the expression and characterization of the genes obtained from these environments is challenging due to effort required to identify appropriate expression hosts and/or conditions for protein purification and characterization.

Enzymatic hydrolysis of lignocellulose is currently one of the most expensive steps in processes for biofuel production. Identifying and/or engineering glycoside hydrolases (GHs) with improved enzymatic properties is a major research challenge in this effort. To this end we have been using a metagenomics approach interrogating a switchgrass-adapted compost microbial community, we identified genes in this community that putatively encode enzymes with diverse activities, including endoxylanase, b-xylosidase, and a-arabinofuranosidase.

In order to validate the metagenomic approach for finding new biocatalysts these ORFs have been cloned, expressed, and assayed for various hemicellulase activities which can then be prioritized for diverse biofuel process conditions. We have begun characterizing the genes that are well-behaved in E. coli, focusing on properties important for the process of biomass hydrolysis, such as thermostability, pH dependence, and ionic liquid tolerance.  Enzymes with promising properties will become the starting points for protein engineering towards the goal of developing a suite of enzymes suitable for efficient hydrolysis of lignocellulosic biomass.