Rapid and discrete isolation of oxygen-evolving His-tagged photosystem II core complex from Chlamydomonas reinhardtii by Ni2+ affinity column chromatography

被引：50

作者：

Sugiura, M ^{[1
]}

Inoue, Y ^{[1
]}

Minagawa, J ^{[1
]}

机构：

[1] RIKEN, Inst Phys & Chem Res, Photosynth Res Lab, Wako, Saitama 3510198, Japan

来源：

FEBS LETTERS | 1998年 / 426卷 / 01期

基金：

日本科学技术振兴机构;

关键词：

photosystem II; psbD; Chlamydomonas reinhardtii; His-tag; Ni2+-chelate chromatography;

D O I：

10.1016/S0014-5793(98)00328-7

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

We have developed a simple and rapid procedure to isolate an oxygen-evolving photosystem II (PS II) core complex from Chlamydomonas reinhardtii. A His-tag made of sis consecutive histidine residues was genetically attached at the carboxy terminus of D2 protein to create a metal binding site on the PS Il supramolecular complex, The recombinant cells producing the His-tagged variant of D2 protein grew photo-autotrophically as well as the wild-type cells. Characterization of the oxygen evolution and the thermoluminescence properties revealed that the His-tagging did not affect the functional integrity of the PS II reaction center. A PS Il core complex was isolated from the detergent-solubilized thylakoids of the recombinant cells in 4 h by a single one-step Ni2+ affinity column chromatography. This preparation consists of D1, D2, CP43, CP47, 33 kDa, and a few low molecular weight proteins, and retains a high rate of oxygen-evolving activity (=1000 mu mol/mg Chl/h), (C) 1998 Federation of European Biochemical Societies.

引用

页码：140 / 144

页数：5

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