马铃薯胃蛋白酶抑制剂的纯化、特性及抑制机理探讨

Purification,Characterization and Reacting Mechanism of Pepsin Inhibitor from Potato

从马铃薯中纯化一种胃蛋白酶抑制剂并对其特性及抑制机理进行探讨。利用醇沉、超滤、DEAE-52和Mono Q离子交换层析法从马铃薯中纯化得到一种主要成分是蛋白质的单亚基胃蛋白酶抑制剂(PPI),相对分子质量为16 100。该抑制剂热稳定性良好,对胃蛋白酶有很强的抑制作用,半抑制浓度IC50值为26.67μg/mL,抑制类型属于非竞争和竞争混合型抑制模式。据圆二色光谱(CD)研究胃蛋白酶抑制剂与胃蛋白酶作用前后的构象变化推测抑制剂以类似线形分子的形式覆盖到靶酶活性中心附近的区域上,从而阻止酶的活性中心与底物接触。

The purpose of this study is to purify a kind of pepsin inhibitor from potato and to study on its characteristics and reacting mechanism. Isolating and purifying the product by alcohol precipitation, Ultrafiltration, DEAE-52 and Mono Q ion exchange chromatography. It was identified to be a protein and showed a single subunit. The relative molecular weight was 16 100. The purified inhibitor had a good thermal stability, and specific inhibition to pepsin. The IC50 to pepsin was 26. 67 μg/mL. The inhibitory type was the complex of non-competitive and competitive inhibitory mode. According to circular dichroism （CD） study of the conformational changes of pepsin inhibitor, pepsin and its complex people might speculate that the inhibitor in the form of a similar linear molecule covered to the enzyme active site, thereby preventing the enzyme active site and substrate contact.