Journal Article

Properties of <i>Chlamydomonas</i> Photosystem II Core Complex with a His-Tag at the C-Terminus of the D2 Protein

Miwa Sugiura, Jun Minagawa and Yorinao Inoue

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 40, issue 3, pages 311-318
Published in print January 1999 | ISSN: 0032-0781
Published online January 1999 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/oxfordjournals.pcp.a029543
Properties of Chlamydomonas Photosystem II Core Complex with a His-Tag at the C-Terminus of the D2 Protein

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  • Plant Sciences and Forestry

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A His-tagged PSII core complex was purified from recombinant Chlamydomonas reinhardtii D2-H thylakoids by single-step Ni2+-affinity column chromatography and its properties were partially characterized in terms of their PSII functions and chemical compositions. The PSII core complex that has a His-tag extension at the C-terminus of the D2 protein evolved oxygen at a high rate of 2,400 μmol (mg Chl)−1h−1 at the optimum pH of 6.5 with ferricyanide and 2,6-dichlorobenzoquinone as electron acceptors in the presence of Ca2+ as an essential cofactor, and approximately 90% of the activity was blocked by 10 μM DCMU. The core complex exhibited the thermoluminescence Q-band but not the B-band regardless of the presence or absence of DCMU, although both bands were observed in the His-tagged thylakoids. The core complex was free from PSI and contained one YD, Tyr 160 of the D2 protein, four Mn atoms, two cytochrome b-559, about 46 Chl a molecules, and probably one QA, the primary acceptor quinone of PSII. It was inferred from these results that His-tagging at the C-terminus of the D2 protein does not affect the functional and structural integrity of the PSII core complex, and that the ‘His-tag strategy’ is highly useful for biochemical, physicochemical, and structural studies of Chlamydomonas PSII.

Keywords: Chlamydomonas reinhardtii; D2 protein; His-tag; Oxygen evolution; PSII core complex; Purification

Journal Article.  0 words. 

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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