An innovative approach of priming lignocellulosics with lytic polysaccharide mono-oxygenases prior to saccharification with glycosyl hydrolases can economize second generation ethanol process

Two Lytic polysaccharide Mono-Oxygenases (LPMOs), non-modular (PMO_08942) and modular (PMO_07920), from thermotolerant fungus Aspergillus terreus 9DR cloned and expressed in Pichia pastoris X33 and purified to homogeneity using ion-exchange chromatography were found to be of similar to 29 and similar to 40 kDa, respectively. Both LPMOs were optimally active at 50 degrees C; PMO_08942 was active under acidic condition (pH 5.0) and PMO_07920 at pH 7.0. Modular LPMO (PMO_07920) tethered to CBM-1 was found to be versatile as it showed appreciable activity on complex polysaccharide (both cellulose and xylans) as compared to non-modular (PMO_08942). The t(1/2) of PMO_08942 (similar to 192 h, pH 5.0) and PMO_0792 (similar to 192 h, pH 7.0) at 50 degrees C, suggests highly stable nature of these LPMOs. Fluorescently tagged modular AA9 was studied microscopically to understand interaction with pretreated biomass. Priming of biomass for up to 6 h with LPMOs prior to initiating hydrolysis with core cellulase enzyme resulted in significantly higher saccharification.