Probe-Based Functional Profiling of Lignocellulose Degrading Microbes

Lignocellulosic biomass has great promise as a highly abundant and renewable source for the production of biofuels. However, the recalcitrant nature of lignocellulose toward hydrolysis into soluble sugars remains a significant challenge to harnessing the potential of this source of bioenergy. Microbial glycoside hydrolases and oxidases play a dominant role in the biochemical conversion of lignocellulosic biomass to high-value biofuels. However, the microbes and/or the enzymes they produce are often intolerant of chemical treatments, high temperatures, and acidic conditions often used for the pretreatment of lignocellulosic material.

We employ activity-based chemical probes to readily characterize and monitor glycoside hydrolase function in diverse microbes – including microbial communities and co-cultures, fungi, anaerobic and aerobic bacteria, and archaea – by LC-MS based proteomics and fluorescent imaging. We demonstrate a unique strength of the probe approach to rapidly identify optimal conditions for maximum enzymatic lignocellulose deconstruction, without requiring numerous cultures. By doing so, we rapidly identify lignocellulolytic enzymes tolerant to extreme temperatures and pHs, ionic liquids, alcohols and other chemicals used in pretreatment methodologies. Importantly, the functional probe approach permits the measurement of individual enzyme activities, not the conglomerate of a microbial protein mix. Finally, the functional probe approach is easy to apply to nearly any lignocellulose-degrading organism, and facilitates research advances toward biofuel development.