Heu M S, Kim H R, Pyeun J H
Department of Nutrition and Food Science, National Fisheries University of Pusan, Korea.
Comp Biochem Physiol B Biochem Mol Biol. 1995 Nov;112(3):557-67. doi: 10.1016/0305-0491(95)00111-5.
The molecular weights of trypsin and chymotrypsin purified from anchovy viscera were estimated to be 25.6 and 26.1 Kda, respectively, by SDS-PAGE. Both enzymes had their maximal activity at pH 9.0 and 45 degrees C for casein and at pH 8.0 and 45 degrees C for synthetic substrates. Trypsin hydrolyzed at the position of Arg22 and Lys29, and chymotrypsin did at the position of Phe1, Tyr16, Phe24, Phe25, and Tyr26 of insulin beta-chain. The K'm and kcat of trypsin were 50 microM and 1.84 microM-1 min-1 toward N-benzoyl-L-arginine-p-nitroanilide (BAPNA) and those of chymotrypsin were 89 microM and 10.0 microM-1min-1 toward N-succinyl-(Ala)2-Pro-Phe-p-nitroanilide. The activation energy of trypsin and chymotrypsin were estimated to be 14 Kcal/mol toward N-benzoyl-L-arginine-p-nitroanilide and 6.5 Kcal/mol toward benzoyl-L-tyrosine ethyl ester.
通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）估计，从鳀鱼内脏中纯化得到的胰蛋白酶和胰凝乳蛋白酶的分子量分别为25.6 kDa和26.1 kDa。两种酶对于酪蛋白在pH 9.0和45℃时具有最大活性，对于合成底物在pH 8.0和45℃时具有最大活性。胰蛋白酶在胰岛素β链的Arg22和Lys29位置进行水解，而胰凝乳蛋白酶在胰岛素β链的Phe1、Tyr16、Phe24、Phe25和Tyr26位置进行水解。胰蛋白酶对N-苯甲酰-L-精氨酸对硝基苯胺（BAPNA）的米氏常数（K'm）和催化常数（kcat）分别为50 μM和1.84 μM⁻¹ min⁻¹，胰凝乳蛋白酶对N-琥珀酰-（丙氨酸）₂-脯氨酸-苯丙氨酸对硝基苯胺的K'm和kcat分别为89 μM和10.0 μM⁻¹min⁻¹。胰蛋白酶和胰凝乳蛋白酶对N-苯甲酰-L-精氨酸对硝基苯胺的活化能估计为14千卡/摩尔，对苯甲酰-L-酪氨酸乙酯的活化能估计为6.5千卡/摩尔。
