A continuous assay of acetate kinase activity: measurement of inorganic phosphate release generated by hydroxylaminolysis of acetyl phosphate.
A continuous assay of acetate kinase activity: measurement of inorganic phosphate release generated by hydroxylaminolysis of acetyl phosphate.
复制标题
乙酸激酶活性的连续测定：测量乙酰磷酸羟氨解产生的无机磷酸盐释放。
DOI：
10.1016/j.bioorg.2007.12.002
复制
发表时间：
2008
影响因子：
5.1
通讯作者：
Sanders,DavidA
中科院分区：
文献类型：
--
作者：
Mukhopadhyay,Soma;Hasson,MiriamS;Sanders,DavidA
乙酸激酶（Acetate kinase，ASKHA）是原核生物碳和能量代谢的核心酶，属于磷酸转移酶超家族。近年来，人们对乙酸激酶和相关羧酸激酶的结构和生物化学进行了广泛的研究。野生型和突变型酶的动力学特性的分析已受到阻碍的性质，目前的测定乙酸激酶活性。这些测定具有不连续或不敏感或利用干扰活性测量的化合物的缺点。我们已经开发了一种新的连续测定，依赖于嘌呤核苷磷酸化酶为基础的光谱测量的羟基氨解的乙酸激酶反应，乙酰磷酸的产物之一产生的无机磷酸盐。该测定能够确定海栖热袍菌乙酸激酶的动力学参数，其表明乙酸的Km比先前公布的低。
Acetate kinase, a member of the ASKHA (Acetate and Sugar Kinases, Hsp70, Actin) phosphotransferase superfamily is a central enzyme in prokaryotic carbon and energy metabolism. Recently extensive structural and biochemical studies of acetate kinase and related carboxylate kinases have been conducted. Analysis of the kinetic properties of wild-type and mutant enzymes has been impeded by the nature of the current assays for acetate kinase activity. These assays have the disadvantages of being either discontinuous or insensitive or of utilizing compounds that interfere with activity measurements. We have developed a novel continuous assay that depends on the purine nucleoside phosphorylase-based spectroscopic measurement of the inorganic phosphate generated by hydroxylaminolysis of one of the products of the acetate kinase reaction, acetyl phosphate. This assay has enabled a determination of the kinetic parameters of the Thermotoga maritima acetate kinase that indicates a lower Kmfor acetate than previously published.