2000 Fiscal Year Final Research Report Summary
Analysis of the genes for porphyrin biosynthetic pathway in higher plants utilizing the light-sensitive mutants of Escherichia coli
| Project/Area Number |
11480201
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| Research Category |
Grant-in-Aid for Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Molecular biology
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
INOKUCHI Hachiro Dept.of Biophys., Gard.Sch.Sci.Kyoto Univ. (Professor), 大学院・理学研究科, 教授 (20028195)
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| Co-Investigator(Kenkyū-buntansha) |
NAKAHIGASHI Kenji Dept.of Biophys., Gard.Sch.Sci.Kyoto Univ.(Assistant Professor), 大学院・理学研究科, 助手 (70322740)
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| Project Period (FY) |
1999 – 2000
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| Keywords | rice (Oryza sativa) / soybean (Glycine max) / hemF gene of E.coli / cucumber (Cucumis sativus) / thymidylate synthase / spinach (Spinacia oleracea) / ferrochelatase / protoporphyrinogen IX oxidase |
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| Research Abstract |
1) By taking advantage of the ability of such a cDNA to complement the thymine auxotrophy of a mutant of Escherichia coli, a cDNA clone for thymidylate synthase (TS) of rice (Oryza sativa L.indica) was isolated. [Kanjo ＆ Inokuchi, Plant Physiol. (Gene Register), 120, 634 (1999)]
2) A cDNA clone for protoporphyrinogen IX oxidase of soybean (Glycine max was isolated by functional complementation against the poor growth of BT3 delta hemG : : Km^R cells. Nucleotide sequence analysis of the cDNA insert revealed that the insert was 1745 bp in length. It included a single open reading frame that encoded a protein consisting of 502 amino acid residues. The deduced amino acid sequence of PPOX from soybean was 67 % homologous to that of mitochondrial PPOX from tobacco. It shares some amino acid residues at the N-terminus speculated to be a mitochondrial translocation signal. [Kanjo et al., Plant Physiol. (Gene Register), 121, 1384 (1999)]
3) We examined poessible alternate pathways for the oxidati
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on of protoporphyrinogen IX to protoporphyrin IX, by isolating and investigating E.coli mutants that can still grow normally when the hemG gene is disrupted. It was suggested that the E.coli aerobic coproporphyrinogen oxidase has an intrinsic capacity to oxidize not only coproporphyrinogen III but also protoporhyrinogen IX.[Narita et al., Mol. Gen. (Genet., 261, 1012-1020 (1999)]
4) A putative mature region of a cucumber (Cucumis sativus) ferrochelatase cDNA (hemH) was overexpressed in E.coli, purified to homogenity and examined its enzymatic properties. Immunolocalization of the enzyme examined with antibody demonstrated that the ferrochelatase was present in both hypocotyls and roots but hardly in cotyledons of cucumber. [Suzuki et al., Plant Cell Physiol., 41, 192-199 (2000)]
5) We reported the first cloning of spinach (Spinacia oleracea) plastidal PPOX cDNA and analysis of the N-terminal sequence in mature plastidal PPOX purified from spinach chloroplast and discussed the mechanism of the transport of plastidal PPOX into chloroplasts. [Che et al., Plant Physiol., 124, 59-70 (2000)] Less
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