Recombinant thermo-alkali-stable endoglucanase of Myceliopthora thermophila BJA (rMt-egl): Biochemical characteristics and applicability in enzymatic saccharification of agro-residues

Codon adaptation index (CAI) of a 1263 bp long endoglucanase encoding gene from the thermophilic mould Myceliopthora thermophile BJA has been improved from 0.44 to 0.76 by in vitro gene synthesis. The codon optimized endoglucanase gene (Mt-egl) has been constitutively expressed in Pichia pastoris under the regulation of GAP promoter. Recombinant endoglucanase (rMt-egl), purified by size exclusion chromatography, has been confirmed to be a monomeric protein of ∼47 kDa. rMt-egl is optimally active at pH 10 and 50 °C, displaying stability in broad pH and temperature ranges, with a t1/2 of 60 and 15 min at 90 and 100 °C, respectively. This retained ∼70% of activity after 3 h incubation at pH 5-12. The Km, Vmax, kcat and kcat/Km of rMt-egl were 5 mg mL−1, 20 μmoles min−1 mg−1, 1.02 × 103 s−1 and 204 s−1 mg−1 mL−1, respectively. Homology modeling and bioinformatics analysis confirmed catalytically important role of glutamate 234 and 344. rMt-egl released high amounts of reducing sugars from wheat bran and corn cobs (421 and 382 mg g−1), thus making it a useful biocatalyst for producing bioethanol and fine chemicals from agro-residues.