Enhancing the Enzymatic Activity of the Multifunctional β-Glycosyl Hydrolase (Cel44C-Man26A_P558) from Paenibacillus polymyxa GS01 Using DNA Shuffling

Abstract

We previously reported that the truncated Cel44C-Man26AP558 β-glycosyl hydrolase protein exhibits multifunctional activities, including cellulase, xylanase, and lichenase. DNA shuffling of the truncated Cel44C-Man26AP558 enzyme was performed to enhance the enzymatic activity of the multifunctional β-glycosyl hydrolase. Two mutant enzymes, M2Cel44C-Man26AP558 that carries one mutation (P438A) and M21Cel44C-Man26AP558 that carries two mutations (A273T and P438A) were obtained. The enzymatic activity of the M21Cel44C-Man26AP558 double mutant was lower than enzymatic activity of the single mutant (M2Cel44C-Man26AP558). However, both mutants displayed the enhancements in their enzyme activities that were ≈1.3-to 2.2-fold higher than the original enzymatic activity in Cel44C-Man26AP558. In particular, the mutant M2Cel44C-Man26AP558 exhibited an approximate 1.5-to 2.2-fold increase in the cellulase, xylanase, and lichenase activities in comparison with the control (Cel44C-Man26AP558). The optimum cellulase, linchenase, and xylanase activities of β-glycosyl hydrolase were observed at pH 7.0, pH 7.0 and pH 6.0, respectively. These results, therefore, suggest that the amino acid residue Ala438 plays important roles in the enhancement of the activity of multifunctional β-glycosyl hydrolase.