純化並定性斑節蝦Penaeus japonicus 肝胰?中β-半乳糖??的活化蛋白

Abstract

利用DE-52陰離子交換管柱，Con A親和力管柱及高效率液相層析逆相碳四管柱（h.p.l.c.RP-C4）可自無脊椎動物斑節蝦肝胰臟中純化出β-半乳糖??的活化蛋白（β-galactosidase activator protein)。此活化蛋白?小分子量（15 kDa)，熱安定（heat stable）的醣蛋白（glycoprotein)，比活性（specific activity)?406 units/mg，較同實驗中由人類胎盤純化的活化蛋白比活性(800 units/mg）低。 在pH4.6的分析緩衝液中，活化蛋白有最佳活性；pH值高於5或低於3.6都不適合活化蛋白的作用。以不同性質（head group電性）介面活性劑（detergent）測試結果指出：在陰離子介面活性劑（anionic detergent）牛磺膽酸鹽（sodium taurocholate）下，活化蛋白活性較強；且最適濃度?0.01％。 本實驗也發現：斑節蝦活化蛋白能加強斑節蝦及jack bean β-半乳糖??的活性；而對α葡萄糖??，β-葡萄糖??沒有作用。其性質與人類saposin B相近，具有專一性（specificity）但無物種間專一性（species specificity）。另外，活化蛋白不影響β-半乳糖??的熱安定性（thermal stability)。

A heat stable, small (apparant molecular mass 15 kDa) glycoprotein was purified as beta-galactosidase activator protein from the hepatopancreas of shrimp Penaeus japonicus with a specific activity of 406 units / mg by using a combination of DE-52 anionic exchange chromatography, Con A chromatography and RP-C4 h.p.l.c. This activator protein stimulates beta-galactosidase activity in a pH range between 3.6 to 5 and has an optimal pH at 4.6. Treatment with various detergents reveals that under anionic detergents such as sodium taurocholate, activator can exert its activity better, and we recommended a concentration of 0.01 %. In our experiment, we found that this activator protein enhances both shrimp and jack bean beta-galactosidase but not alpha-glucosidase or beta-glucosidase which is characteristic for human saposin B. In addition, activator protein shows no effects on the thermal stability of beta-galactosidase.