Heterologous expression and characterization of gene from in

Abstract

?-Mannanase catalyzes endo-wise hydrolysis of the backbone of mannan and heteromannan, which are abundant in the cell wall structure of ungerminated leguminous seeds. The mature ?-mannanase originated from Bacillus subtilis was expressed in Pichia pastoris, a methylotrophic yeast, using the leader peptide sequence of Saccharomyces cerevisiae ?-factor. The cultivation of ?-mannanase expressing Pichia pastoris yields up to 1.8 g/L protein. In the supernatant the activity of the 40 kDa-total mannanase attained a level of 1102.0 IU/mL. The properties of the ?-mannanase were characterized. Optimum pH and temperature for the recombinant enzyme were 5.5 and 50°C respectively. The enzyme was stable at pH 5.0–10.0 and maintained over 30% original activity after incubating at 70°C for 30 min.

Keywords

β-mannanase, Pichia pastoris, expression