(560e) One-Step Production of Lactate and Isobutanol From Pretreated Biomass by Recombinant Cellulolytic Bacillus Subtilis

Consolidated bioprocessing (CBP) is low-cost cellulosic biomass bioconversion by integrating cellulase production, cellulose hydrolysis, and sugar fermentation into a single step. Although significant efforts had been made to create recombinant cellulolytic microorganisms, real recombinant cellulolytic microorganism that can produce sufficient secretory active cellulases, hydrolyze cellulose, and utilize soluble sugars for supporting cell growth and cellulase synthesis without any other organic nutrient is not available yet. We demonstrated that over-expression of Bacillus subtilis endoglucanase BsCel5 by more than 30-fold enables non-cellulose-utilizing Bacillus subtilis to grow on amorphous cellulose and pretreated lignocellulosic biomass as the sole carbon source without addition of costly organic nutrients. After directed evolution and screening on amorphous cellulose, both the expression/secretion level and specific activity of Cel5 and Cel9 had been increased successfully. An advanced recombinant cellulolytic Bacillus subtilis strain has been constructed with engineered cellulase and the genes knock out/knock in. Besides these, we also achieved the co-expression and secretion of family 5, 9 and 48 cellulases in a single B. subtilis strain. The one-step production of lactate and isobutanol from cellulose and pretreated biomass in the minimal medium without adding any organic nutrient had been demonstrated. We are reconstituting in vivo mini-cellulosome on the cell surface of B. subtilis. The recombinant cellulolytic B. subtilis would be an ultra-low-cost platform for producing biofuels and other value-added products from non-food biomass.