OPTA对乳酸脱氢酶的抑制动力学  

KINETICS OF INACTIVATION OF LDH DURING IRREVERSIBLE MODIFICATION BY OPTA

作　　者：杨晓[1] 吴海斌[2] 赵康源[2] 

机构地区：[1]北京协和医院,北京100730 [2]中国科学院生物物理研究所生物大分子国家重点实验室,北京100101

出　　处：《生物物理学报》1993年第3期362-368,共7页Acta Biophysica Sinica

基　　金：国家自然科学基金39270170部分资助

摘　　要：邹承鲁建立的酶活性不可逆改变动力学理论已为实验所验证,它不仅适用于单底物酶的抑制和激活的动力学研究,而且也适用于双底物酶反应系统.但在双底物酶反应系统中,底物和酶的结合方式有四种机制,即随机机制、有序机制、强制有序机制和乒乓机制,迄今为止这一动力学方法仅对随机机制的肌酸激酶进行了实验研究.而其它机制的实验研究尚未见诸报道.我们选用了有序机制的乳酸脱氢酶(LDH),用邻苯二甲醛(OPTA)为抑制剂对该酶的抑制过程进行了实验研究.结果表明,OPTA对该酶的抑制为不可逆抑制.其产物生成与时间的关系曲线符合邹氏方程:[P]=[P]_x(1-e^(-A[OPTA]).由ln([P]_x-[P])对t作图为一直线,表明它的抑制作用为单相动力学过程,抑制剂与酶的结合为一步反应.由直线的斜率A[OPTA]对[OPTA]作图为一过原点的直线.说明表观速度常数A与OPTA的浓度无关.OPTA与酶的结合为非络合型.测得的OPTA与EE-NADH结合的微观速度常数分别为:K(?)=49.6(mmol L)^(-1)min,(?)=2.31(mmol L)min^(-1)(?).明显小于(?)的事实表明.NADH对失活有明显的保护作用.OPTA是一个竞争性的不可逆抑制剂.用传统的方法测得的(?)为42.5(mmol L)min^(-1).与邹氏方法测得的结果非常接近.Kinetics of inactivation of LDH by OPTA has been studied by following the substrate reaction in the presence of the inactivator. The microscopic constants for the reaction of the inactivator with the free-enzyme and with the enzyme - substrate complexes were determined. Results showed that with respect to substrate, NADH, the inactivator, OPTA, is competitive. The formation of the enzyme -substrate complex protects against the inactivation by OPTA. The inactivation kinetics is monophasic with OPTA. and the results also suggested that OPTA is a noncomplexing inhibiton for LDH. The present paper proves that the theory of the kinetics of substrate reaction during irreversible modification of enzyme activity can also be applied to enzyme reactions involving two substrates forming a ternary complex by an ordered sequence.

关 键 词：动力学 不可逆抑制 乳酸脱氢酶 

分 类 号：Q61[生物学—生物物理学]