IMMOBILIZED ARTIFICIAL MEMBRANE CHROMATOGRAPHY - RAPID PURIFICATION OF FUNCTIONAL MEMBRANE-PROTEINS

被引:59
作者
PIDGEON, C
STEVENS, J
OTTO, S
JEFCOATE, C
MARCUS, C
机构
[1] PURDUE UNIV,SCH PHARM & PHARMACAL SCI,DEPT PHARMACOL & TOXICOL,W LAFAYETTE,IN 47907
[2] PURDUE UNIV,SCH PHARM & PHARMACAL SCI,DEPT MED CHEM & PHARMACOGNOSY,W LAFAYETTE,IN 47907
[3] UNIV WISCONSIN,SCH MED,DEPT PHARMACOL,MADISON,WI 53706
关键词
D O I
10.1016/0003-2697(91)90164-O
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
A solid-phase membrane mimetic system, denoted as immobilized artificial membranes (IAM), has been developed and utilized as a novel high-performance liquid chromatography (HPLC) matrix for the first step in the rapid purification of functional membrane proteins. IAM phases consist of monolayers of amphiphilic membrane lipid molecules covalently bonded to a rigid silica particle. These monolayers of lipids have proved remarkably effective for the chromatography of biomolecules. Several cytochrome P450 isozymes, an extremely important family of hydrophobic membrane proteins with a labile heme catalytic center, have been partially purified in functional conformations from rat liver, kidney, and adrenal microsomes on IAM supports. Functionality of purified P450 and P450 reductase has been demonstrated by optical difference spectroscopy, by carbon monoxide binding, and by reconstitution of enzymatic activity in vitro. Other membrane proteins, including rat liver plasma membrane NADH oxidase and ferricyanide oxidoreductase have also been partially purified by IAM HPLC. The methods for purification of these proteins are described. © 1991.
引用
收藏
页码:163 / 173
页数:11
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