College of Biological Engineering, The Key Laboratory of Science and Technology for Aquaculture and Food Safety, Jimei University, Jimei, Xiamen, 361021, China.
Comp Biochem Physiol B Biochem Mol Biol. 2011 Apr;158(4):259-65. doi: 10.1016/j.cbpb.2010.12.003. Epub 2010 Dec 15.
Three pepsinogens (PG1, PG2, PG3) were highly purified from the stomach of Japanese seabass (Lateolabrax japonicus) by ammonium sulfate fractionation, DEAE-Sephacel anion exchange column chromatography and Sephacryl S-200 gel-filtration. Two dimensional polyacrylamide gel electrophoresis (2D-PAGE) analysis revealed that the molecular masses of the three PGs were 35, 37, and 34kDa, and their isoelectric points were 5.3, 5.1, and 4.7, respectively. Zymography analysis showed that the three pepsinogens had different mobilities and enzymatic activities under native conditions. Pepsinogens converted into their active form pepsins under pH 2.0 by one-step pathway or stepwise pathway. All three pepsins were completely inhibited by pepstatin A, a typical aspartic proteinase inhibitor. The N-terminal amino acid sequences of the three pepsinogens were determined to the 30th, 30th and 28th amino acid residue and those of their corresponding active form pepsins were also determined to the 19th, 18th and 20th amino acid residue, respectively. All amino acid sequences of Japanese seabass PGs revealed high identities to reported fish and mammalian pepsinogens. The effective digestion of fish and shrimp muscular proteins by pepsins indicated their physiological function in the degradation of food proteins.
三种胃蛋白酶原（PG1、PG2、PG3）通过硫酸铵分级沉淀、DEAE-葡聚糖阴离子交换柱层析和 Sephacryl S-200 凝胶过滤从日本鲈鱼（Lateolabrax japonicus）的胃中高度纯化。二维聚丙烯酰胺凝胶电泳（2D-PAGE）分析表明，这三种 PG 的分子量分别为 35、37 和 34kDa，等电点分别为 5.3、5.1 和 4.7。酶谱分析表明，三种胃蛋白酶原在天然条件下具有不同的迁移率和酶活性。胃蛋白酶原在 pH 2.0 下通过一步途径或逐步途径转化为其活性形式胃蛋白酶。三种胃蛋白酶均被典型的天冬氨酸蛋白酶抑制剂胃蛋白酶抑制剂 A 完全抑制。三种胃蛋白酶原的 N 端氨基酸序列分别测定到第 30、30 和 28 位氨基酸残基，其相应的活性形式胃蛋白酶也分别测定到第 19、18 和 20 位氨基酸残基。日本鲈鱼 PG 的所有氨基酸序列与报道的鱼类和哺乳动物胃蛋白酶原具有高度的同源性。胃蛋白酶有效消化鱼类和虾类肌肉蛋白表明它们在食物蛋白降解中的生理功能。
