| Project/Area Number |
08457633
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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 |
応用薬理学・医療系薬学
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| Research Institution | Gunma University School of Medicine |
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Principal Investigator |
KOHAMA Kazuhiro Gunma University, School of Medicine, Department of Pharmacology, Professor, 医学部, 教授 (30101116)
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| Co-Investigator(Kenkyū-buntansha) |
中村 彰男 群馬大学, 医学部, 助手 (30282388)
ISHIKAWA Ryoki Gunma University, Department of Pharmacology, Assistant Professor, 医学部, 助手 (20212863)
OKAGAKI Tsuyoshi Gunma University, School of Medicine, Department of Pharmacology, Lecturer, 医学部, 講師 (80185412)
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| Project Period (FY) |
1996 – 1997
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| Project Status |
Completed (Fiscal Year 1997)
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| Budget Amount *help |
¥7,200,000 (Direct Cost: ¥7,200,000)
Fiscal Year 1997: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 1996: ¥5,800,000 (Direct Cost: ¥5,800,000)
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| Keywords | smooth muscle / myosin light chain kinase / calcium / regulation / recombinant protein / calmodulin / actin / myosin / カルモジュリン / アクチン / カルシウムイオン / アクトミオシン |
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| Research Abstract |
Myosin light chain kinase (MLCK) plays a central role in regulating the actin-myosin interaction of smooth muscle. MLCK phosphorylates the light chain of myosin in the presence of Ca^<2+> and calmodulin (CaM) thereby activating myosin so that it can interact with actin. Besides this kinase activity, MLCK shows i) actin-binding activity that can assemble actin filaments into their bundles and ii) myosin-binding activity that can form myosin filaments. To localize the actin- and myosin-binding activities in the MLCK molecule and to examine their possible role in regulating the actin myosin interaction, we expressed various fragments of cDNA encoding MLCK in Escherichia coli as recombinant proteins. We found that MLCK consists of an N-terminal actin-binging domain, a central kinase domain, and a C-terminal myosin-binding domain. The Met^1-Pro^<41> sequence is responsible for Ca^<2+>/CaM-sensitive binding to actin. This binding site exerts an inhibitory effect on the actin-myosin interaction only when myosin is phosphorylated. MLCK binds to myosin at the C-terminal domain, the sequence of which is identical to telokin, an abundant myosin-binding protein in smooth muscle cells. This domain itself has no regulatory role in the interaction. However, the interaction was stimulated when this domain was extended to include the sequence known to regulate the activity of the kinase domain. The stimulation was observed only when myosin was unphosphorylated.
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