糖对长白猪精液NAGaseⅡ的抑制机理

Inhibition mechanism of saccharide on NAGaseⅡfrom sperm of landrace

以长白猪精液为材料,采用经典的纯化方法获得N-乙酰-β-D-氨基葡萄糖苷酶(EC 3.3.1.52,NAGase)的2种同工酶,命名为:NAGaseⅠ和NAGaseⅡ。其中NAGaseⅡ的比活力为358.21U/mg,纯化倍数为6.37倍。以海藻糖、D-甘露糖、蔗糖、葡萄糖为效应物,研究其对该酶活性的影响。结果表明:在特定浓度范围内,海藻糖对酶活力基本没有影响;D-甘露糖、蔗糖、葡萄糖对该酶均有一定的抑制作用;D-甘露糖对酶的抑制作用表现为非竞争型,抑制常数KI为11.94mmo/L;蔗糖对酶的抑制作用表现为竞争型,抑制常数KI为0.56mmol/L;葡萄糖对酶的抑制作用表现为混合型,抑制常数KI和KIS分别为0.35mol/L和64.29mmol/L。

There were two kinds of N-acetyl-β-D-glucosaminidase（EC3. 2.1. 52, NAGase） in the Landrace semen,who were named as NAGase I and NAGase Ⅱ. NAGase Ⅱ was purified from the Landrace semen by ammonium sulfate fractionation, chromatography on DEAE-32 and Sephadex G-100. The specific activity of the enzyme was 358.21 U/mg with purification by 6.37-fold. The influence of the enzyme activity by Trehalose,D-mannose,Sucrose,Glucose were found that Trehalose had no effects on the activity of the enzyme lD-mannose,Sucrose and Glucose showed various degrees of inhibitory effects on the enzyme activity,D-Mannose was a non-competitive inhibitor with the inhibition constants of free enzyme （KI） was 11.94 mmol/L. Sucrose was a competi- tive inhibitor with the inhibition constants of free enzyme （K1） was 0.56 mmol/L. Glucose was a mixed-type inhibitor with the inhibition constants of free enzyme （K1） and enzyme-substrate complex （Kts） were 0.49 mmol/L and 64.29 mmol/L,respectively.