Hwa-Ja Ryu, Eun-Seong Seo, Hee-Kyoung Kang, Young-Min Kim, Doman Kim
Abstract
A gene encoding human intestinal maltase (HMA) was successfully expressed in Pichia pastoris under the control of the methanol-induced alcohol oxidase (AOX1) promoter. The secreted recombinant HMA fused with a His6-tag was produced (150 U/L) and was easily purified from culture supernatants in a 3-step diafiltration, ultrafiltration, and affinity column chromatography protocol. The specific activity of the purified HMA was 16.8 U/mg. Endoglycosidase H digestion of the protein showed that the recombinant HMA was N-glycosylated. The purified HMA was maximally active at pH 6.5 and stable (≥90%) up to 65°C. The kinetic parameters K m and V max were 3.3±0.25 mM maltose and 61.9±2 U/mg, respectively.
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Keywords: α-glucosidase,maltase,diabetes,Pichia pastoris,expression in yeast.
