(123f) Inhibitory Effect of Cellobionic Acid on Cellobiohydrolase and Cellobiose Dehydrogenase

The development of microbial fermentation platforms for the production of organic acids has gained interest in the last decade due to the reliability and cost-effectiveness of such processes.  In recent years, carboxylic acids have emerged as specialty acids due to their unique physiochemical properties.  Cellulosic biomass, which is available at low cost and in widespread abundance, is a potential substrate for the bio-production of carboxylic acids.  We have engineeredNeurospora crassa for the production of cellobionic acid (CBA) directly from cellulose with exogenous laccase and redox mediator addition.  In this system, cellulases convert cellulose to cellobiose, cellobiose dehydrogenase (CDH) oxidizes cellobiose to CBA, and CDH is continuously regenerated by laccase and a redox mediator.  While inhibition of cellulases by cellobiose has been extensively studied, cellobionic acid inhibition of cellulases has not received much attention, despite the discovery that oxidative enzymes produced by cellulolytic organisms generate aldonic acids from cellulosic substrates.  In this study, we investigate the inhibition of cellobiohydrolase I by cellobiose and CBA using p-nitrophenyl-β-D-lactopyranoside as a model cellulosic substrate.  Additionally, CBA inhibition of CDH is evaluated.  The kinetic properties and mechanism of CBA inhibition are studied and compared to that of cellobiose inhibition.