Heterologous Expression of Pectin Methylesterase Inhibitor and its Application in Reducing Methanol in Fruit Wine
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Abstract:
The pectin methylesterase inhibitor (PMEI) gene, cloned from the kiwi fruit, was linked to the expression vector pPICzαA through double digestion using EcoRI/XbaI. It was then transferred into Pichia pastoris GS115 by electro-transformation to obtain the positive strain GS115/pPICzαA-PMEI. After the PMEI expressed by GS115/pPICzαA-PMEI was concentrated and purified, it was applied in fruit wine fermentation. The results show that the recombinant P. pastoris expression strain GS115/pPICzαA-PMEI was successfully constructed, and the PMEI expression level was 35.38 mg/L. When PMEI was applied in fruit wine fermentation, the methanol in orange wine decreased by 47.54% from 189.75~99.55 mg/L. Similarly, the methanol content of apple wine dropped by 21.52% from 118.15~91.20 mg/L, whereas the methanol content of grape wine did not change significantly and remained below 20.00 mg/L. These results indicate that PMEI has a significant methanol reduction effect on oranges that are rich in pectin and endogenous pectinase. The minimum inhibitory concentration of PMEI for optimal orange wine fermentation was 8.84 mg/L and optimum temperature was 18 ℃. Under these conditions, the methanol content of orange wine reduced by 66.40%, to 89.10 mg/L. These findings provide a novel technique for producing low-methanol fruit wine by applying PMEI.
