The redox regulation of melatonin synthesis in circadian rhythm
| Project/Area Number |
16590049
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Biological pharmacy
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| Research Institution | SHUJITSU UNIVERSITY |
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Principal Investigator |
TSUBOI Seiji Shujitsu University, Faculty of Pharmaceutical Science, Professor, 薬学部, 教授 (50172052)
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| Co-Investigator(Kenkyū-buntansha) |
YATSUSHIRO Syouki 就実大学, 薬学部, 助手 (90399155)
MORIYAMA Yoshinori Okayama University, Faculty of Pharmaceutical Science, Professor, 大学院・医歯薬学総合研究科, 教授 (10150658)
OHTSUKA Masato Okayama University, Advanced Science Research Center, 自然生命科学支援センター, 助手 (30243489)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2005: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2004: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Keywords | melatonin / serotonin N-acetyltransferase / redox regulation / circadian rhythm / disulfide bonde / glutathione / pinal gland |
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| Research Abstract |
Serotonin N-acetyltransferase (EC.2.3.1.87)(AA-NAT) is a melatonin rhythm-generating enzyme in pineal glands. To establish a melatonin rhythm, AA-NAT activity is precisely regulated through several signaling pathways. Here we show novel regulation of AA-NAT activity, in which an intramolecular disulfide bond may function as a switch for the catalysis. Recombinant-AA-NAT activity was irreversibly inhibited by N-ethylmaleimide(NEM) in an acetyl CoA-protectable manner. Oxidized glutathione or dissolved oxygen reversibly inhibited AA-NAT in an acetyl CoA-protectable manner. To identify the cysteine residues responsible for the inhibition, AA-NAT was first oxidized with dissolved oxygen, treated with NEM, reduced with dithiothreitol, and then labeled with [^<14>C]-NEM. Cys61 and Cys177 were specifically labeled in an acetyl CoA-protectable manner. The AA-NAT with the Cys61 to Ala and Cys177 to Ala double substitutions(Cys61Ala/Cys177Ala-AA-NAT) was fully active but did not exhibit sensitivity to either oxidation or NEM, while the AA-NATs with only the single substitutions retained about 40 % of these sensitivities. An intramolecular disulfide bond between Cys61 and Cys177 formed upon oxidation and cleaved upon reduction was identified. Furthermore, Cys61Ala/Cys177Ala-AA-NAT expressed in COS7 cells was relatively insensitive to H2O2-evoked oxidative stress, while wild type AA-NAT was strongly inhibited under the same conditions. These results indicate that the formation and cleavage of the disulfide bond between Cys61 and Cys177 produce the active and inactive states of AA-NAT. It is possible that intracellular redox conditions regulate AA-NAT activity through switching via an intramolecular disulfide bridge.
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Report
(3 results)
Research Products
(8 results)
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[Journal Article] Change in the Redox State of Glutathione Regulates Differentation of Tracheary Elements in Zinnia Cells and Arabidopsis Roots2005
Author(s)
Henmi, K., Demura, T., Tsuboi, S., Fukada, H., Iwabuchi, M., Ogawa, K.
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Journal Title
Plant Cell Physiol. 46(11)
Pages: 1757-1765
Description
「研究成果報告書概要(和文)」より
Related Report
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[Journal Article] Change in the Redox State of Glutathione Regulates Differentation of Tracheary Elements in Zinnia Cells and Arabidopsis Roots2005
Author(s)
Henmi, K., Demura, T., Tsuboi, S., Fujada, H., Iwabuchi, M., Ogawa, K.
-
Journal Title
Plant Cell Physiol. 46(11)
Pages: 1757-1765
Description
「研究成果報告書概要(欧文)」より
Related Report
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