Journal Article

Significance of Zinc in a Regulatory Protein, CCM1, Which Regulates the Carbon-Concentrating Mechanism in <i>Chlamydomonas reinhardtii</i>

Tsutomu Kohinata, Haruku Nishino and Hideya Fukuzawa

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 49, issue 2, pages 273-283
Published in print February 2008 | ISSN: 0032-0781
Published online February 2008 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pcn003
Significance of Zinc in a Regulatory Protein, CCM1, Which Regulates the Carbon-Concentrating Mechanism in Chlamydomonas reinhardtii

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In conditions with the poor availability of inorganic carbon (CO2 and HCO3 : Ci) for photosynthesis, aquatic photosynthetic organisms induce active Ci uptake systems that allow accumulation of Ci within the cell, the so-called carbon-concentrating mechanism (CCM). In a unicellular green alga, Chlamydomonas reinhardtii, a regulatory factor CCM1 is indispensable for the regulation of the CCM by sensing CO2 availability. CCM1 has two putative zinc-binding domains with several conserved cysteine and histidine residues in its N-terminal region. To determine whether the domains actually bind zinc atoms, the N-terminal parts of CCM1 were expressed as glutathione S-transferase fusion proteins and subjected to atomic absorption spectrometry. It was found that 1 mol of zinc is bound to 1 mol of amino acid regions 1–71 and 72–101 of CCM1, respectively. In the case of the site-directed mutant proteins, H54Y, C77V and C80V, the zinc-binding ability was lost. Physiological analyses of the transgenic Chlamydomonas cells harboring a mutated Ccm1 gene revealed that amino acid residues such as C36, C41, H54, C77, C80, H90 and C93 were indispensable for induction of the CCM in response to Ci-limiting stress conditions. Size exclusion chromatography followed by immunoblot analyses indicated that CCM1 is present as a protein complex of approximately 290–580 kDa independent of Ci availability.

Keywords: Carbonic anhydrase — Carbon-concentrating mechanism — Carbon dioxide — Photosynthesis — Signal transduction — Zinc

Journal Article.  6795 words.  Illustrated.

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

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