Influence of culture aeration on the cellulase activity of Thermobifida fusca

Abstract

Currently, one of the hurdles hindering efficient production of cellulosic biofuel is the recalcitrant nature of cellulose to hydrolysis. A wide variety of cellulase enzymes are found natively in microorganisms that can potentially be used to effectively hydrolyze cellulose to fermentable sugars. In this study, phenomenological and mechanistic parameters affecting cellulase activity were studied using the moderately thermophilic, aerobic, and cellulolytic microorganism Thermobifida fusca. Two major sets of experiments were conducted to (1) study the mechanistic differences in growth in a flask compared to a bioreactor and (2) study the cell culture parameters influencing cellulase activity using a series of bioreactor experiments. Specific cellulase and specific endoglucanase activities were found to be higher in the bioreactor as compared to flask growth. Measurements of messenger RNA transcript levels of 18 cellulase-related genes and intracellular ATP levels indicated that measured enzyme activity was likely more influenced by post-transcriptional energetics rather than transcriptional regulation. By delineating the effects of culture aeration and stir speed using a bioreactor, it was found that cellulase activity increased with increasing aeration and increasing stir speeds (highest K(l)a) with a tradeoff of decreased cellular growth at the highest stir speeds tested (400 rpm). Overall, these results allude to a connection between aeration and oxidative respiration that lead to increased ATP allowing for increased cellulase synthesis as the primary constraint on overall cellulase activity.