Klomklao Sappasith, Benjakul Soottawat, Visessanguan Wonnop, Kishimura Hideki, Simpson Benjamin K
Department of Food Science and Technology, Faculty of Technology and Community Development, Thaksin University, Phattalung Campus, Phattalung, 93110, Thailand.
J Agric Food Chem. 2007 May 30;55(11):4548-53. doi: 10.1021/jf063319x. Epub 2007 May 1.
Trypsin from the pyloric ceca of Atlantic bonito (Sarda sarda) was purified and characterized with respect to its purity; molecular weight; sensitivity to temperature, pH, and inhibition; and N-terminal sequence. The purified trypsin had a molecular weight of 29 kDa as per sodium dodecyl sulfate polyacrylamide gel electrophoresis, and optimal activity was observed at pH 9 and 65 degrees C with BAPNA as a substrate. The enzyme was stable to heat treatment up to 50 degrees C and within the pH range of 7-12. It was stabilized by calcium ions, but its activity was strongly inhibited by soybean trypsin inhibitor, N-p-tosyl-L-lysine chloromethyl ketone, and phenyl methyl sulfonyl fluoride. The enzyme exhibited a progressive decrease in activity with increasing NaCl concentration (0-30%). The N-terminal 20 amino acid residues of Atlantic bonito trypsin were determined as IVGGYECQAHSQPWQPVLNS and were homologous with other trypsins.
对大西洋狐鲣（Sarda sarda）幽门盲囊中提取的胰蛋白酶进行了纯化，并对其纯度、分子量、对温度、pH值和抑制剂的敏感性以及N端序列进行了表征。根据十二烷基硫酸钠聚丙烯酰胺凝胶电泳结果，纯化后的胰蛋白酶分子量为29 kDa，以BAPNA为底物时，在pH 9和65℃条件下观察到最佳活性。该酶在高达50℃的热处理条件下以及pH值7至12范围内稳定。它可被钙离子稳定，但活性受到大豆胰蛋白酶抑制剂、N-对甲苯磺酰-L-赖氨酸氯甲基酮和苯甲基磺酰氟的强烈抑制。随着NaCl浓度的增加（0 - 30%），该酶的活性逐渐降低。大西洋狐鲣胰蛋白酶的N端20个氨基酸残基被确定为IVGGYECQAHSQPWQPVLNS，与其他胰蛋白酶具有同源性。
