2000 Fiscal Year Final Research Report Summary
Roles and specific functions of homologous enzymes involved in biogenesis of active-form sulfur and active-form selenium
| Project/Area Number |
11480179
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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 |
Functional biochemistry
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
ESAKI Nobuyoshi Kyoto University, Institute for Chemical Research, Professor, 化学研究所, 教授 (50135597)
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| Co-Investigator(Kenkyū-buntansha) |
KURIHARA Tatsuo Kyoto University, Institute for Chemical Research, Instructor, 化学研究所, 助手 (70243087)
YOSHIMURA Tohru Kyoto University, Institute for Chemical Research, Associate Professor, 化学研究所, 助教授 (70182821)
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| Project Period (FY) |
1999 – 2000
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| Keywords | selenocysteine lyase / cysteine desulfurase / selenophosphate / selenium / iron-sulfur cluster / sulfur |
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| Research Abstract |
Mechanism and physiological role of cysteine desulfurase which decomposes cysteine into alanine and sulfur and its homolog, selenocysteine lyase, were studied. Two cysteine desulfurases from a cyanobacterium were used to generate an iron-sulfur cluster in an iron-sulfur protein. The enzymes efficiently converted apo-ferredoxin into [2Fe-2S] ferredoxin by using L-cysteine as a sulfur source. Selenocysteine lyase from Escherichia coli was employed to synthesize selenophosphate in vitro. Selenophosphate was efficiently produced in the presence of selenophosphate synthetase, selenocysteine, and ATP.Three gene-disrupted E.coli strains, in which a gene for each cysteine desulfurase homolog was disrupted, were constructed to investigate role of each enzyme. Growth rate of the iscS-disruptant was significantly decreased on a LB plate medium. The iscS-disruptant was transformed with plasmids containing genes for CSD, CsdB, IscS and their mutant enzymes, and growth-rates of the transformatnts were investigated on various solid media. Introduction of csdB into the iscS-disruptant recover the growth-rate of the strain to the level as the wild-type strain. The result suggests that the over-produced CsdB can be a back-up of IscS in vivo. On the other hand, expression of CSD did not complement the growth of the iscS-disruptant. Thus, the in vivo functions of CSD and CsdB are different from each other, although they show about 50% amino acid identity.
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