Heterologous Expression and Enzyme Properties of β-mannanase from Trichoderma reesei in Pichia pastoris

Endo-1,4-β-mannanase from Trichoderma reesei is an important member of glycoside hydrolase family 5. To investigate the characteristics of the enzyme, the gene of the enzyme was obtained by RT-PCR and connected to the plasmid named pPIC9 k to construct a recombinant vector. The recombinant vector was converted into Pichia pastoris to express the target protein. The purified protein was used to investigate the characteristics of the recombinant enzyme. The optimal pH for the recombinant mannanase to action was 6.0, and the optimal temperature was 80 °C. What’s more, the enzyme had a high alkali resistance and a relatively poor acid resistance. It showed higher thermostability at 60 °C than 70 °C. K+, Zn2+, Fe3+, Ca2+ and Mg2+ promoted the activity of the enzyme in different degree, while Fe2+, Cu2+, Li2+ and EDTA had a strong inhibitory effect on the activity of the recombinant mannanase. Ni2+ and Co2+ have little influence on the activity of recombinant enzyme. Through calculation, Km was 0.5 mg/mL, Vmax was 253.8 IU/mg.