An Investigation of the Secretome Composition of Mafic-2001 and the Optimization of the Mafic-2001 Enzyme Cocktail to Enhance the Saccharification Efficacy of Chinese Distillers' Grains.

The efficient degradation of lignocellulose is essential for valorizing agricultural waste and reducing environmental pollution. An efficient degradation process requires an enzyme cocktail capable of comprehensively deconstructing lignocellulosic components. In this study, the secretome of Mafic-2001 induced by rice straw was examined, and the enzymatic composition and reaction conditions of were optimized. Mafic-2001 secreted an enzyme cocktail that included ligninolytic enzymes, cellulases, and hemicellulases. However, the relative abundances of endoglucanase (EG) and β-glucosidase (βG) were only 64.37% and 10.69%, respectively, compared with the relative abundance of cellobiohydrolase, which indicated a critical bottleneck in degradation efficiency. To overcome this limitation, the recombinant enzymes rEG1 and rβG1 were expressed in X-33. A functionally enhanced enzyme cocktail (rEG1-rβG1-Mafic-2001 = 0.05:0.09:0.86) was developed via a mixture design to achieve a reducing sugar yield of 2.77 mg/mL from Chinese distillers' grains (CDGs). Structural analyses revealed that the optimized enzyme cocktail disrupted the reticulated fiber architecture of CDGs and attenuated the characteristic Fourier-transform infrared spectroscopy peaks of lignin, cellulose, and hemicellulose. This study elucidates the synergistic lignocellulose deconstruction mechanism of Mafic-2001 and establishes a precision enzyme-supplementation strategy for efficient CDG bioconversion, providing a scalable platform for the valorization of lignocellulosic biomass.