Engineering E. Coli Whole-Cell Catalysts for Consolidated Production of Lactic Acid From Cellodextrin

            Engineering E. coli whole-cell catalysts for consolidated production of lactic acid from cellodextrin

Charles Rutter, Nimmy Mathew, Rachel Chen

School of Chemical and Biomolecular Engineering, Georgia Institute of Technology

Reduction of production cost for bio-based fuels and chemicals is an important engineering objective yet to be achieved. Consolidated bioprocessing is one of the approaches being pursued by many laboratories around the globe and offers an opportunity for significant cost reduction by combining cellulase production, cellulose depolymerization, and product synthesis in a single step. Engineering microbial catalyst capable of performing these three tasks is, however, challenging. As a consequence, consolidated processing remains largely laboratory adventures, despite significant progresses in the past few years.

In order to develop a system to be used for consolidated bioprocessing of cellulose, expression and secretion of β-glucosidase and cellodextrinase must be achieved.  This presentation describes the use of a heterologous cellodextrinase to enable E. coli cells to grow on cellodextrins and cellobiose.  This enzyme has been shown to release glucose from cello-oligomers across a range of degrees of polymerization with highest activity on cellotetraose.  Additionally, we have shown that this heterologous lipoprotein is properly transolcated, acylated, and inner membrane associated in the E. coli host.  Finally, periplasmic displayed enzyme conferred the cells  the ability to metabolize cellodextrins and ferment them to lactic acid at nearly 75% of theoretical yield.

            This study marks the first attempt at using an inner membrane anchored lipoprotein as an enzyme for degradation of substrates located in the extracellular medium. The ability of this enzyme to hydrolyze extracellular without being translocated across the outer membrane helps to reduce the energetic requirements of this strain for use in consolidated bioprocesses.  Additionally, this study demonstrates that heterologous lipoproteins can be properly modified post-translationally by E. coli and localized to the periplasmic space.