Toyota Eiko, Iyaguchi Daisuke, Sekizaki Haruo, Itoh Kunihiko, Tanizawa Kazutaka
Faculty of Pharmaceutical Sciences, Health Sciences University of HokkaidoIshikari-Tobetsu, Hokkaido 061-0293, Japan.
Biol Pharm Bull. 2007 Sep;30(9):1648-52. doi: 10.1248/bpb.30.1648.
Three isoforms of anionic chum salmon trypsin (ST-1, ST-2, and ST-3) were purified from the pyloric caeca of chum salmon (Oncorhynchus keta). The molecular weights of the three isoforms were about 24 kDa as determined by SDS-PAGE. The isoelectric points of ST-1, ST-2, and ST-3 were 5.8, 5.4, and 5.6, respectively. The apparent K(m) values of two isoforms (ST-1 and ST-2) for BAPA (benzoyl-L-arginine-p-nitroanilide) hydrolysis at 5, 15, 25 and 35 degrees C were slightly higher than that of the main isoform ST-3, depending on temperature. The turnover numbers, k(cat), of ST-1 and ST-2 were about twice as high as that of ST-3. Consequently, the catalytic efficiencies (k(cat)/K(m)) of ST-1 and ST-2 were more efficient than ST-3. There were marked differences in both apparent K(m) and k(cat) values of three anionic chum salmon trypsins as compared to bovine cationic trypsin. K(m) values of all chum salmon trypsins were approximately 10 times lower than those of bovine trypsin, depending on the temperature. The k(cat) values of all chum salmon trypsins were about 2- to 5-fold higher than those of bovine trypsin; therefore, the catalytic efficiencies (k(cat)/K(m)) of chum salmon trypsin were 20- to 40-fold more efficient than those of bovine trypsin. On the other hand, k(cat)/K(m) values of ST-1 for TAME (tosyl-L-arginine methyl ester) hydrolysis were lower than those of bovine trypsin, whereas k(cat)/K(m) values of ST-2 and ST-3 were comparable to those of bovine trypsin, depending on the temperature.
从大麻哈鱼（Oncorhynchus keta）的幽门盲囊中纯化出了三种阴离子型大麻哈鱼胰蛋白酶同工型（ST-1、ST-2和ST-3）。通过SDS-PAGE测定，这三种同工型的分子量约为24 kDa。ST-1、ST-2和ST-3的等电点分别为5.8、5.4和5.6。两种同工型（ST-1和ST-2）在5、15、25和35摄氏度下催化BAPA（苯甲酰-L-精氨酸对硝基苯胺）水解的表观K(m)值略高于主要同工型ST-3，具体取决于温度。ST-1和ST-2的转换数k(cat)约为ST-3的两倍。因此，ST-1和ST-2的催化效率（k(cat)/K(m)）比ST-3更高。与牛阳离子胰蛋白酶相比，三种阴离子型大麻哈鱼胰蛋白酶的表观K(m)值和k(cat)值均存在显著差异。取决于温度，所有大麻哈鱼胰蛋白酶的K(m)值约比牛胰蛋白酶低10倍。所有大麻哈鱼胰蛋白酶的k(cat)值比牛胰蛋白酶高约2至5倍；因此，大麻哈鱼胰蛋白酶的催化效率（k(cat)/K(m)）比牛胰蛋白酶高20至40倍。另一方面，取决于温度，ST-1催化TAME（甲苯磺酰-L-精氨酸甲酯）水解的k(cat)/K(m)值低于牛胰蛋白酶，而ST-2和ST-3的k(cat)/K(m)值与牛胰蛋白酶相当。
