大鼠肝脏丝氨酸羟甲基转移酶的亲和纯化及特性研究

Affinity purification and characterization of serine hydroxymethyltransferases from rat liver.

作者信息

Masuda T, Sakamoto M, Nishizaki I, Hayashi H, Yamamoto M, Wada H

出版信息

J Biochem. 1987 Mar;101(3):643-52. doi: 10.1093/jb/101.3.643.

DOI:10.1093/jb/101.3.643

Abstract

A rapid and simple method was developed for the purification of serine hydroxymethyltransferases [EC 2.1.2.1]. The procedure involved ammonium sulfate precipitation, DEAE-cellulose column chromatography and affinity chromatography on an L-adsorbent. Through this procedure the cytosolic enzyme (s-SHMT) was purified 1,650-fold, and the mitochondrial enzyme (m-SHMT) 1,730-fold, with a yield of more than 30% in both cases. Both preparations gave a single band with a Mr of 54,000 on SDS-PAGE. The native enzymes both contained 4 mol of pyridoxal phosphate/mol of enzyme, and showed a Mr value of 220,000 on gel filtration, indicating a tetrameric structure. Several other properties of the enzymes were also studied.

摘要

开发了一种快速简便的方法用于纯化丝氨酸羟甲基转移酶[EC 2.1.2.1]。该方法包括硫酸铵沉淀、DEAE-纤维素柱色谱和在L吸附剂上的亲和色谱。通过该方法，胞质酶（s-SHMT）纯化了1650倍，线粒体酶（m-SHMT）纯化了1730倍，两种情况下产率均超过30%。两种制剂在SDS-PAGE上均呈现一条Mr为54,000的条带。天然酶均含有4摩尔磷酸吡哆醛/摩尔酶，在凝胶过滤中显示Mr值为220,000，表明为四聚体结构。还研究了酶的其他几个性质。