两种纤溶酶的酶学性质比较与溶栓机理分析  

Characterization and Fibrinolytic Mechanism of Two Fibrinolytic Enzymes

作　　者：华颖[1] 沈国华[1] 刘大群[1] 

机构地区：[1]浙江省农业科学院食品科学研究所,杭州310021

出　　处：《中国食品学报》2012年第10期47-55,共9页Journal of Chinese Institute Of Food Science and Technology

基　　金：浙江省自然科学基金资助(No.Y3090482)

摘　　要：对芽孢杆菌(Bacillus sp.nov.SK006)发酵产生的两种纤溶酶(SPFE-1和SPFE-2)的酶学性质和溶栓机理进行分析比较,并研究其酶促反应动力学参数。试验结果表明:SPFE-1和SPFE-2具有较好的耐热性和pH稳定性;Zn2+能促进SPFE-1的纤溶活力,但强烈抑制SPFE-2;PMSF对SPFE-1h具有强烈的抑制作用;SPFE-1和SPFE-2都可直接降解纤维蛋白,也可激活纤溶酶原,从而间接降解纤维蛋白;SPFE-1最先降解纤维蛋白原的β链,然后是α链,很难降解γ链;SPFE-2顺次降解纤维蛋白原的α、β和γ链;SPFE-1和SPFE-2有较强的酰胺水解活性,最适底物为N-Succ-Ala-Ala-Pro-Phe-pNA;其作用于最佳底物的特征常数:Km=0.6/0.51mmol/L;kcat=712/154.25/s;kcat/Km=1.19×106/3.02×105L/(mol·s)。Two fibrinolytic enzymes（SPFE-1and SPFE-2） were produced from Bacillus sp. nov. SK006. The enzymes displayed optimal activities at different temperatures and pH values. Zinc ion stimulated the enzyme activity of SPFE-1 whereas caused effective activity inhibition of SPFE-2. The fibrinolytic activity of SPFE-1 was strongly inhibited by PMSF, and moderately inhibited by EDTA as well as PCMB. EDTA and 2-mercaptoethanol affected SPFE-2 significantly. During the degradation of fibrinogen by SPFE-1, B β-chains of fibrinogen were cleaved first, followed by slower release of the γ-chains. The A α-subunit was resistant to the enzyme digestion. SPFE-2 was more effective with the A α-subunit, but less sensitive towards γ-chains. Both enzymes exhibited higher affinity towards N-Succ-Ala-Ala-Pro-Phe-pNA. The synthetic substrates were effectively hydrolyzed with amidolytic activity of 158.18 and 107.47 nmol/min/mL, Km of 0.60 and 0.51 mmol/L, as well as kcat/Km of 1.19×106 and 3.02×105 L/（mol·s）（SPFE-1and SPFE-2）, respectively. Both enzyme were able to degrade fibrin clots either by forming active plasmin from plasminogen or by direct fibrinolysis. N-terminal amino acid sequences of the enzymes were found to be AQSVPYEQPHLSQ.

关 键 词：芽孢杆菌 纤溶酶SPFE-1和SPFE-2酶学性质 纤溶机理 反应动力学 

分 类 号：TS201.25[轻工技术与工程—食品科学]