1996 Fiscal Year Final Research Report Summary
Molecular Organization of Photosystem II Reaction Center
| Project/Area Number |
06404003
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
植物生理
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| Research Institution | Okayama University |
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Principal Investigator |
SATOH Kimiyuki Okayama University, Faculty of Science, Department of Biology, Professor, 理学部, 教授 (10032822)
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| Co-Investigator(Kenkyū-buntansha) |
INAGAKI Noritoshi Naitonal Institute for Basic Biology, Research Associate, 基礎生物学研究所, 助手 (50221776)
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| Project Period (FY) |
1994 – 1996
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| Keywords | Photosynthesis / Photosystem II / Reaction Center / Synechocystis / Mutagenesis / Chlamydomonas / P-680 / Processing Protease |
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| Research Abstract |
Dynamic as well as static organization of molecules in the photosystem II reaction center was analyzed in this project.
I.Structure of isolated photosystem II reaction center
(1) Both subunit stoichiometry and pigment stoichiometry of the isolated photosystem II reaction center were re-evaluated in this study using refined techniques.
(2) beta-Carotenes in the purified photosystem II reaction center were selectively extracted with diethyl ether containing varied amount of water and the organization and interaction of pigments in the reaction center were analyzed using materials with different pigment contents.
(3) The contribution of a pheophytin molecule in inactive branch of electron transport system in the absorption spectrum of photosystem II reaction center was analyzed by fluorescence excitation spectroscopy as well as by absorption, linear dichroism and magnetic circular dichroism spectra. The results were discussed in the light of current understanding of the molecular organization
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in photosystem II raction center.
II.Dynamic aspects
(1) The light-regulated synthesis of the D1 subunit of photosystem II reaction center was analyzed using isolated pea chloroplasts. In that study we have found that the translation of D1 protein is regulated at the specific steps of polypeptide elongation by a redox factor (s) that is activated by the reduction via photosystem I.We also have succeeded in partially purifying this factor.
(2) The enzyme involved in the C-terminal processing of precursor D1 protein was purified from sonicated extracts of thylakoids, by a method that includes chromatography on QAE-anion exchange, hydroxylapatite, Cu-chelating affinity and gel-filtration columns. Based on the amino acid sequence data of the purified protease, a cDNA clone encoding the enzyme was identified and sequenced, from a spinach green leaf cDNA library. By these analyzes, the full-length transcript was established to consist of 1906 nucleotides and a poly (A) tail, containing an open reading frame (ORF) corresponding to a protein with 539 amino acid residues. By comparing the amino acid sequence of the purified protease with that deduced from nucleotide sequence of the cDNA clones, the enzyme was shown to be furnished with an extra N-terminal extension consisted of 150 amino acids which is characteristic of both a transit peptide and a signal sequence. The mechanism of enzymatic catalysis was analyzed using enzymes over-expressed in E.coli. Less
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