Acan N L, Ozer N
Department of Biochemistry, Faculty of Medicine, Hacettepe University, Ankara, Turkey.
J Enzyme Inhib. 2001 Nov;16(5):457-64. doi: 10.1080/14756360109162395.
Human erythrocyte pyruvate kinase was modified with bromopyruvate and the kinetic behavior of the modified enzyme was investigated. When the enzyme was modified with bromopyruvate in the absence of adenosine-5'-diphosphate, phosphoenolpyruvate or fructose-1,6-diphosphate the inactivation followed a pseudo first-order kinetics. The inactivation rate constant, ks, was 1.84 +/- 0.15 min(-1). Kd of the bromopyruvate-enzyme complex was 0.14 +/- 0.03 mM. The presence of adenosine-5'-diphosphate, phosphoenolpyruvate or fructose-1,6-diphosphate in the modification medium or the presence of fructose-1,6-diphosphate in the assay medium resulted in deviation of the inactivation kinetics from pseudo first-order. Phosphoenolpyruvate was better than adenosine-5'-diphosphate for protection against bromopyruvate modification whereas fructose-1,6-diphosphate was ineffective. The modified enzyme showed negative cooperativity in the presence of fructose-1,6-diphosphate whereas in the absence of it no activity was detected.
用溴丙酮酸修饰人红细胞丙酮酸激酶，并研究修饰后酶的动力学行为。当在不存在腺苷-5'-二磷酸、磷酸烯醇丙酮酸或果糖-1,6-二磷酸的情况下用溴丙酮酸修饰该酶时，失活遵循假一级动力学。失活速率常数ks为1.84±0.15 min⁻¹。溴丙酮酸-酶复合物的Kd为0.14±0.03 mM。修饰介质中存在腺苷-5'-二磷酸、磷酸烯醇丙酮酸或果糖-1,6-二磷酸，或测定介质中存在果糖-1,6-二磷酸，导致失活动力学偏离假一级。磷酸烯醇丙酮酸在防止溴丙酮酸修饰方面比腺苷-5'-二磷酸更好，而果糖-1,6-二磷酸无效。修饰后的酶在存在果糖-1,6-二磷酸时表现出负协同性，而在不存在果糖-1,6-二磷酸时未检测到活性。
