College of Food Science and Technology, Nanjing Agricultural University, Nanjing, Jiangsu, China.
J Sci Food Agric. 2011 Oct;91(13):2437-42. doi: 10.1002/jsfa.4484. Epub 2011 Jun 23.
Proteases have become an essential part of the modern food and feed industry, being incorporated in a large and diversified range of products for human and animal consumption. The objective of this study was to purify and characterise a protease from wheat germ.
After purification a single protease of molecular weight 61-63 kDa (determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis) was obtained. The purified protease had optimal activity at 50 °C and maintained its activity completely after incubation at 30 °C for 30 min, while over 47% of the activity was lost after incubation at 80 °C for 30 min. The purified protease had optimal activity and maintained maximum stability at pH 5.5, while the activity decreased after incubation for 30 min at other pH values. The protease was inhibited by Mg(2+), Mn(2+), Ba(2+) and iodoacetic acid and stimulated by Li(+), Ca(2+), Cu(2+), β-mercaptoethanol and dithiothreitol, while Zn(2+), L-cysteine and glutathione had no significant effect on its activity. At pH 5.5 the enzyme had a K(m) of 0.562 mg mL(-1) with casein as substrate and showed higher affinity to casein than to bovine serum albumin, ovalbumin and gelatin.
The purified enzyme from wheat germ was identified as a cysteine protease.
蛋白酶已成为现代食品和饲料工业的重要组成部分，广泛应用于人类和动物食用的各种产品中。本研究的目的是从麦胚中纯化和鉴定一种蛋白酶。
经纯化后得到一种分子量为 61-63 kDa 的单一蛋白酶（通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳测定）。该纯化蛋白酶在 50°C 时具有最佳活性，在 30°C 下孵育 30 分钟后仍保持其全部活性，而在 80°C 下孵育 30 分钟后则损失超过 47%的活性。该纯化蛋白酶在 pH5.5 时具有最佳活性和最大稳定性，而在其他 pH 值下孵育 30 分钟后其活性降低。该蛋白酶被 Mg(2+)、Mn(2+)、Ba(2+)和碘乙酸抑制，被 Li(+)、Ca(2+)、Cu(2+)、β-巯基乙醇和二硫苏糖醇刺激，而 Zn(2+)、L-半胱氨酸和谷胱甘肽对其活性没有显著影响。在 pH5.5 时，该酶对酪蛋白的 K(m)为 0.562mg mL(-1)，对酪蛋白的亲和力高于牛血清白蛋白、卵清蛋白和明胶。
从麦胚中纯化的酶被鉴定为一种半胱氨酸蛋白酶。
