Production of biohydrogen from hydrolyzed bagasse with thermally preheated sludge

Production of biohydrogen from dark fermentation is an interesting alternative to producing renewable fuels because of its low cost and various usable substrates. Cellulosic content in plentiful bagasse residue is an economically feasible feedstock for biohydrogen production. A statistical experimental design was applied to identify the optimal condition for biohydrogen production from enzymatically hydrolyzed bagasse with 60-min preheated seed sludge. The bagasse substrate was first heated at 100 °C for 2 h and was then hydrolyzed with cellulase. Culture of the pretreated bagasse at 55 °C provided a higher H₂ production performance than that obtained from cultures at 45 °C, 65 °C, 35 °C and 25 °C. On the other hand, the culture at pH 5 resulted in higher H₂ production than the cultures at pH 6, pH 4 and pH 7. The optimal culture condition for the hydrogen production rate was around 56.5 °C and pH 5.2, which was identified using response surface methodology. Moreover, the pretreatment of bagasse under alkaline conditions gave a thirteen-fold increase in H₂ production yield when compared with that from preheatment under neutral condition.     

Characterization of the Biomass Degrading Enzyme GuxA from Acidothermus cellulolyticus

Microbial conversion of biomass relies on a complex combination of enzyme systems promoting synergy to overcome biomass recalcitrance. Some thermophilic bacteria have been shown to exhibit particularly high levels of cellulolytic activity, making them of particular interest for biomass conversion. These bacteria use varying combinations of CAZymes that vary in complexity from a single catalytic domain to large multi-modular and multi-functional architectures to deconstruct biomass. Since the discovery of CelA from Caldicellulosiruptor bescii which was identified as one of the most active cellulase so far identified, the search for efficient multi-modular and multi-functional CAZymes has intensified. One of these candidates, GuxA (previously Acel_0615), was recently shown to exhibit synergy with other CAZymes in C. bescii, leading to a dramatic increase in growth on biomass when expressed in this host. GuxA is a multi-modular and multi-functional enzyme from Acidothermus cellulolyticus whose catalytic domains include a xylanase/endoglucanase GH12 and an exoglucanase GH6, representing a unique combination of these two glycoside hydrolase families in a single CAZyme. These attributes make GuxA of particular interest as a potential candidate for thermophilic industrial enzyme preparations. Here, we present a more complete characterization of GuxA to understand the mechanism of its activity and substrate specificity. In addition, we demonstrate that GuxA exhibits high levels of synergism with E1, a companion endoglucanase from A. cellulolyticus. We also present a crystal structure of one of the GuxA domains and dissect the structural features that might contribute to its thermotolerance.