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

FEBS Letters

Inoue, Yorinao¹  Minagawa, Jun¹  Sugiura, Miwa¹ 

[1] Photosynthesis Research Laboratory, Institute of Physical and Chemical Research (RIKEN), Wako, Saitama 351-0198, Japan

关键词: Photosystem II;    psbD;    Chlamydomonas reinhardtii;    His-tag;    Ni2+-chelate chromatography;    Chl;    chlorophyll;    DCBQ;    dichlorobenzoquinone;    DCMU;    3-(3;    4-dichlorophenyl)-1;    1-dimethyl urea;    DM;    n-dodecyl β-d-maltoside;    LHC;    light-harvesting complex;    PS I;    photosystem I;    PS II;    photosystem II;    PAGE;    polyacrylamide gel electrophoresis;    WT;    wild type;   

DOI  :  10.1016/S0014-5793(98)00328-7

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 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 six consecutive histidine residues was genetically attached at the carboxy terminus of D2 protein to create a metal binding site on the PS II supramolecular complex. The recombinant cells producing the His-tagged variant of D2 protein grew photoautotrophically 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 II 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 μmol/mg Chl/h).

【 授权许可】

Unknown