College of Food Science and Technology, Yunnan Agricultural University, Kunming 650201, Yunnan, China.
Biotechnology and Germplasm Resources Institute, Yunnan Academy of Agricultural Sciences, Kunming 650223, China.
Int J Biol Macromol. 2021 Feb 28;171:539-549. doi: 10.1016/j.ijbiomac.2021.01.026. Epub 2021 Jan 9.
The present study aimed to investigate the biological functions of germinated M. oleifera seed proteins and to identify the identity of milk-clotting proteases. A total of 963 proteins were identified, and those with molecular weights between 10 and 30 kDa were most abundant. The identified proteins were mainly involved in energy-associated catalytic activity and metabolic processes, and carbohydrate and protein metabolisms. The numbers of proteins associated with the hydrolytic and catalytic activities were higher than the matured dry M. oleifera seeds reported previously. Of the identified proteins, proteases were mainly involved in the milk-clotting activity. Especially, a cysteine peptidase with a molecular mass of 17.727 kDa exhibiting hydrolase and peptidase activities was purified and identified. The identified cysteine peptidase was hydrophilic, and its secondary structure consisted of 27.60% alpha helix, 9.20% beta fold, and 63.20% irregular curl; its tertiary structure was also constructed using M. oleifera seed 2S protein as the protein template. The optimal pH and temperature of the purified protease were pH 4.0 and 60 °C, respectively. The protease had high acidic stability and good thermostability, thus could potentially be applied in the dairy industry.
本研究旨在探讨发芽麻疯树种子蛋白的生物学功能，并鉴定凝乳蛋白酶的特性。共鉴定到 963 种蛋白质，其中分子量在 10-30 kDa 之间的蛋白质最为丰富。鉴定到的蛋白质主要参与与能量相关的催化活性和代谢过程，以及碳水化合物和蛋白质代谢。与水解和催化活性相关的蛋白质数量高于之前报道的成熟干麻疯树种子。在所鉴定的蛋白质中，蛋白酶主要参与凝乳活性。特别是，一种分子量为 17.727 kDa 的半胱氨酸肽酶，具有水解酶和肽酶活性，被分离和鉴定。鉴定出的半胱氨酸肽酶具有亲水性，其二级结构由 27.60%的α螺旋、9.20%的β折叠和 63.20%的不规则卷曲组成；其三级结构也使用麻疯树种子 2S 蛋白作为蛋白质模板进行构建。纯化蛋白酶的最适 pH 和温度分别为 pH 4.0 和 60°C。该蛋白酶具有较高的酸性稳定性和良好的热稳定性，因此有可能应用于乳制品行业。
