一种用于黄素激酶的连续荧光测定法。来自埃氏消化链球菌的黄素激酶的性质。

A continuous fluorometric assay for flavokinase. Properties of flavokinase from Peptostreptococcus elsdenii.

作者信息

Mayhew S G, Wassink J H

出版信息

Biochim Biophys Acta. 1977 Jun 10;482(2):341-7. doi: 10.1016/0005-2744(77)90247-9.

PMID:195619

Abstract

A continuous fluorometric assay that utilizes apoflavodoxin as a trapping agent for riboflavin 5'-phosphate (FMN) has been developed for flavokinase (ATP:riboflavin 5'-phosphotransferase, EC 2.7.1.26). Use of this assay is illustrated in a procedure for the partial purification of flavokinase from the strict anaerobe Peptostreptococcus elsdenii. The purified enzyme catalyzed the formation of 8.3 nmol FMN - min-1 - mg-1 at 37 degrees C and had apparent Km values for riboflavin and ATP of 10 and 4.7 micronM, respectively. ATP could be replaced by ADP (22% of the rate observed with ATP) but not by GTP. The enzyme also phosphorylated 5-deaza- and 8-bromoriboflavin with activities of 15 and 70%, respectively, of that with riboflavin; it was inactive with iso riboflavin and deoxyriboflavin.

摘要

已开发出一种连续荧光测定法，该方法利用脱辅基黄素odoxin作为核黄素5'-磷酸（FMN）的捕获剂来测定黄素激酶（ATP：核黄素5'-磷酸转移酶，EC 2.7.1.26）。在从严格厌氧菌埃尔氏消化链球菌中部分纯化黄素激酶的过程中展示了该测定法的应用。纯化后的酶在37℃下催化形成FMN的速率为8.3 nmol·min⁻¹·mg⁻¹，对核黄素和ATP的表观Km值分别为10和4.7 μM。ATP可用ADP替代（为ATP时观察到的速率的22%），但不能用GTP替代。该酶还能使5-脱氮核黄素和8-溴核黄素磷酸化，其活性分别为核黄素的15%和70%；对异核黄素和脱氧核黄素无活性。