Molecular Dissection and Application of an Enzyme-Inhibitor System with Special Reference to a Proteinase That Specifically Decomposes a Hardly Soluble Protein in Food Seeds.

从小麦种子中发现的一种新的半胱氨酸蛋白酶--麦醇溶蛋白酶中重组得到一种酶。它实际上分解，虽然在体外，麦醇溶蛋白作为一个主要的小麦蛋白质，并被发现参与蛋白质水解的储存蛋白在发芽过程中。其次，我们以oryzacystatin cDNA为探针，通过筛选克隆了小麦半胱氨酸蛋白酶抑制剂。由此获得了编码半胱氨酸蛋白酶抑制剂的三个cDNA克隆，并且公开了所有这些半胱氨酸蛋白酶抑制剂都属于所谓的“植物半胱氨酸蛋白酶抑制剂”家族。同时，我们通过与谷胱甘肽S-转移酶（GST）融合，制备了重组麦醇溶蛋白。在弱酸性条件下，GST-前麦醇溶蛋白具有蛋白水解活性，该活性可被市售的卵半胱氨酸蛋白酶抑制剂和谷半胱氨酸蛋白酶抑制剂所强烈抑制，也可被我们重组获得的两种小麦半胱氨酸蛋白酶抑制剂所抑制。所有这些结果表明，小麦半胱氨酸蛋白酶的蛋白质水解和小麦半胱氨酸蛋白酶抑制剂的调节系统的可能性，可以在工业上应用于食品加工面粉品质的改善。

A recombinant enzyme was produced from gliadain we found as a new cysteine proteinase in wheat seeds. It actually decomposed, though in vitro, gliadin as a major wheat protein and was found to be involved in the proteolysis of the storage protein during germination. Next, we cloned wheat cystatins by screening with oryzacystatin cDNA as a probe. Three cDNA clones encoding cystatins were thus obtained and it was disclosed that all these cystatins belong to the so-called "phytocystatin" family. In the mean time, we produced a recombinant species of gliadain by fusion with glutathione S-transferase (GST). The resultant GST-progliadain elicited a proteolytic activity under a weakly acidic condition, which was strongly inhibited by a commercially available preparation of egg cystatin as well as by oryzacystatin.The activity was also inhibited to a similar extent by two wheat cystatins we obtained recombinantly. All these results suggest the possibility that a system of the proteolysis with wheat cysteine proteinases and its regulation with wheat cystatins can be industrially applied to food processing for flour quality improvement.