对异源表达的肉碱棕榈酰转移酶II的动力学、圆二色性和荧光研究。

Kinetic, circular dichroism and fluorescence studies on heterologously expressed carnitine palmitoyltransferase II.

作者信息

Mann W R, Yan B, Dragland C J, Bell P A

机构信息

Metabolic Diseases Department, Sandoz Research Institute, Sandoz Pharmaceuticals Corp., East Hanover, New Jersey 07936-1080, USA.

出版信息

J Enzyme Inhib. 1995;9(4):303-8. doi: 10.3109/14756369509036559.

DOI:10.3109/14756369509036559

PMID:8598540

Abstract

Km estimates for carnitine and palmitoyl-CoA of heterologously expressed rat liver carnitine palmitoyl-transferase-II (rCPT-II) were 950 +/- 27 microM and 34 +/- 6 microM, respectively. Vmax for the enzyme was 1.8 mumol/min/mg purified protein. Consistent with an ordered reaction mechanism in which palmitoyl-CoA binds first, SDZ CPI 975, a reversible carnitine palmitoyltransferase inhibitor containing both carnitine and alkyl moieties, inhibited rCPT-II competitively with carnitine and uncompetitively with palmitoyl-CoA. Substrate-enzyme interactions were examined by circular dichroism (CD) and fluorescence. Both carnitine and palmitoyl-CoA alone induced conformational changes in the enzyme; dissociation constant estimates by CD for carnitine and palmitoyl-CoA were 41 +/- 5 microM and 7 +/- 2 microM, respectively.

摘要

异源表达的大鼠肝脏肉碱棕榈酰转移酶-II（rCPT-II）对肉碱和棕榈酰辅酶A的米氏常数（Km）分别为950±27微摩尔和34±6微摩尔。该酶的最大反应速度（Vmax）为1.8微摩尔/分钟/毫克纯化蛋白。与棕榈酰辅酶A先结合的有序反应机制一致，SDZ CPI 975是一种同时含有肉碱和烷基部分的可逆性肉碱棕榈酰转移酶抑制剂，它对rCPT-II的抑制作用表现为对肉碱是竞争性的，对棕榈酰辅酶A是非竞争性的。通过圆二色性（CD）和荧光对底物与酶的相互作用进行了研究。单独的肉碱和棕榈酰辅酶A都会诱导该酶的构象变化；通过CD估算的肉碱和棕榈酰辅酶A的解离常数分别为41±5微摩尔和7±2微摩尔。