| Project/Area Number |
15K05557
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Research Field |
Bio-related chemistry
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| Research Institution | Fukushima University |
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Principal Investigator |
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| Co-Investigator(Renkei-kenkyūsha) |
MURAYAMA Kazutaka 東北大学, 大学院医工学研究科, 准教授 (40400452)
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| Project Period (FY) |
2015-04-01 – 2018-03-31
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| Project Status |
Completed (Fiscal Year 2017)
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| Budget Amount *help |
¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2017: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
Fiscal Year 2016: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2015: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
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| Keywords | Phospholipase D / ホスホリパーゼD（PLD） / Lysoplasmalogen-PLD / Choline lysoplasmalogen / Lyso-PAF（リゾ型血小板因子） / 基質認識メカニズム / 基質特異性改変 / 構造活性（機能）相関 / リゾプラズマローゲン / ホスホリパーゼD / 部位特異的変異導入 / 速度論解析 / 立体構造予測 / 部位特異的変異解析 / プラズマローゲン |
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| Outline of Final Research Achievements |
Choline lysoplasmalogen-specific phospholipase D (LyPlsCho-PLD) is an enzyme that catalyzes the hydrolytic cleavage of the phosphoester bond of LyPls, releasing choline and lysoplasmenic acid. LyPlsCho-PLD exhibited weak activity toward LysoPAF. Our goal is to elucidate the substrate recognition mechanism of LyPlsCho-PLD. Based on the homology-modeled structure of LyPls-PLD and the docking model with substrate analog, amino acid residues predicted to interact with the head group and sn-1-alkenyl ether chain were substituted by site-directed mutagenesis. The princial conclusions of the study by mutation analysis and kinetics were as follows: 1) M71, N173 and F211 of LyPlsCho-PLD may be involved in the recognition of sn-1 alkenyl ether chain of LyPlsCho. F211 and W282 may take part in substrate binding and reaction products release (=k+2). 2) F211A, I, L, and C variants remarkably altered in the substrate preference (LysoPAF >> LyPlsCho) while maintaining the high activity.
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