大豆胰蛋白酶抑制剂的异源表达与生化特性解析

Heterologous Expression and Biochemical Characterization of Soybean Trypsin Inhibitor

本文对一疑似大豆胰蛋白酶抑制剂(STI;sti)的开放阅读框在毕赤酵母中进行了克隆表达与生化特性分析。结果表明,在摇瓶水平上,重组菌GS115(p PIC-STI)分泌表达了30 mg/L STI;重组STI在(40～80)℃或pH 2.0～11.0孵育1 h后,仍能保持85%以上的抑制活性;K^+、Zn^(2+)和Mg^(2+)对其胰酶抑制活性有明显的激活作用,而Cu^(2+)、Mn^(2+)、Ca^(2+)、Fe^(2+)和Fe^(3+)则有明显的抑制作用;重组STI对胰蛋白酶具有较强的、专一性的和非竞争性的抑制作用,是一种典型的多肽类胰酶抑制剂。良好的酶学性质使STI在食品、医药等行业具有潜在的应用价值。

A putative trypsin inhibitor from Glycine max(namely STI encoded by sti)was expressed in Pichia pastoris and its biochemical characteristics were investigated.Resuts showed that:In shaking-flask fermentation experiments,recombinant yeast GS115(pPIC-STI)secretorily expressed 30mg/L STI.The recombinant STI retained more than 85% of its maximum inhibitory activity after incubation at 40-80℃ or pH 2.0-11.0 for 1h.Its activity was significantly enhanced by K^+,Zn^2+ and Mg^2+,but strongly inhibited by Cu^2+,Mn^2+,Ca^2+,Fe^2+ and Fe^3+.The STI exerted the strongest,specific and non-competitive inhibitory effects toward trypsin,which indicated that STI was a typical peptide-type trypsin inhibitor.All these distinct biochemical properties make STI a good candidate for food and pharmaceutical applications.