Cloning and characterization of a cold-adapted endo-1,5-α-L-arabinanase from Paenibacillus polymyxa and rational design for acidic applicability.

AbnZ1, with optimal pH of 6.0 and optimal temperature of 40 °C, is a cold-adapted endo-1,5-α-L-arabinanase encoded by the gene abnZ1 from Paenibacillus polymyxa Z6. The specific activity of AbnZ1 remained 54.1% of maximum at 5 °C. To apply AbnZ1 in acidic conditions, three basic hsitidine (His) residues, His(48), His(218), and His(297), around the catalytic domain were selected as mutation sites, which were replaced with Asp, Glu, Arg, and Lys, respectively, to yield 12 mutants, H48D/E/R/K, H218D/E/R/K, and H297D/E/R/K. The optimum pH of mutant H218D shifted toward the acidic direction by 0.5 unit, and the relative activity was enhanced from 20.4 to 55.7% at pH 5.0. Furthermore, the specific activity of H218D in optimal conditions was 82.6 U/mg versus that of wild type, 73.4 U/mg, and the K(m) decreased from 11.9 to 7.1 mg/mL. This work provided an arabinanase candidate for juice clarification and pectin extraction.