Expression of a wheat β-1,3-glucanase in Pichia pastoris and its inhibitory effect on fungi commonly associated with wheat kernel

Abstract β -1,3-glucanases, the plant PR-2 family of pathogenesis-related (PR) proteins, can be constitutively expressed and induced in wheat crop to enhance its anti-fungal pathogen defense. This study aimed to investigate the inhibitory effect of wheat β -1,3-glucanase on fungi most commonly associated with wheat kernel. A β -1,3-glucanase from wheat was successfully expressed in Pichia pastoris X-33 and its biochemical and antifungal properties were characterized herein. The molecular weight of recombinant β -1,3-glucanase is approximately 33 kDa. β -1,3-glucanase displays optimal activity at pH 6.5, remaining relatively high at pH 5.5–8.0. The optimal reaction temperature of β -1,3-glucanase is 50 °C, retaining approximately 84.0% residual activity after heat-treated at 50 °C for 1 h. The steady-state kinetic parameters of β -1,3-glucanase against laminarin was determined and the K m and V max were 1.32 ± 0.20 mg/ml and 96.4 ± 4.4 U mg −1 protein, respectively. The inhibitory effect of purified β -1,3-glucanase against the seven fungi commonly associated with wheat kernel was assessed in vitro. β -1,3-glucanase exerted differential inhibitory effects on hyphal growth of Fusarium graminearum , Alternaria sp. , A. glaucus , A. flavus , A. niger , and Penicillium sp. Spore formation and mycelial morphology of Alternaria sp. , A. flavus , and A. niger were significantly affected by β -1,3-glucanase (1U). The present results would help elucidate the mechanism underlying the inhibition of wheat β -1,3-glucanases on pathogens.