Abstract
In the context of avoiding the use of non-renewable energy sources, employing lignocellulosic biomass for ethanol production remains a challenge. Cellulases play an important role in this scenario: they are some of the most important industrial enzymes that can hydrolyze lignocellulose. This study aims to improve on the characterization of a thermostable Aspergillus fumigatus endo-1,4-β-glucanase GH7 (Af-EGL7). To this end, Af-EGL7 was successfully expressed in Pichia pastoris X-33. The kinetic parameters Km and Vmax were estimated and suggested a robust enzyme. The recombinant protein was highly stable within an extreme pH range (3.0–8.0) and was highly thermostable at 55 °C for 72 h. Low Cu2+ concentrations (0.1–1.0 mM) stimulated Af-EGL7 activity up to 117%. Af-EGL7 was tolerant to inhibition by products, such as glucose and cellobiose. Glucose at 50 mM did not inhibit Af-EGL7 activity, whereas 50 mM cellobiose inhibited Af-EGL7 activity by just 35%. Additionally, the Celluclast® 1.5L cocktail supplemented with Af-EGL7 provided improved hydrolysis of sugarcane bagasse “in natura”, sugarcane exploded bagasse (SEB), corncob, rice straw, and bean straw. In conclusion, the novel characterization of Af-EGL7 conducted in this study highlights the extraordinary properties that make Af-EGL7 a promising candidate for industrial applications.
Highlights
Raw material processing generates tons of agricultural and industrial waste every year
The recombinant vector pPICZαA/Af-egl7 was transformed into E. coli DH10β for plasmid propagation
The recombinant plasmid pPICZαA/Af-egl7 was linearized with the restriction enzyme PmeI to enable its integration into the P. pastoris X-33 genome by homologous recombination, and transformed into this strain
Summary
Raw material processing generates tons of agricultural and industrial waste every year. Increased energy consumption, depletion of fossil fuel sources, and the need to abate global warming have drawn special attention to a new generation of renewable energy [1,2,3]. In this scenario, reusing lignocellulosic residues to synthesize chemical compounds and high-value products like biofuels and other green chemicals has emerged as one of the most potential strategies to overcome environmental problems [2,4]. The classic mechanism of substrate degradation by cellulases involves the synergistic action of three types of hydrolytic enzymes: endo-1,4-β-glucanases (EC 3.2.1.4), cellobiohydrolases/exoglucanases (EC 3.2.1.91 and EC 3.2.1.176), and β-glucosidades (EC 3.2.1.21) [8,9]
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