Changes in the Substrate Specificities of Farnesyl Diphosphate Synthase by a Single Amino Acid Substitution

单一氨基酸取代对法尼基二磷酸合酶底物特异性的变化

• 批准号: 12680587
• 负责人: MAKI Yuji
• 金额: $ 2.37万
• 依托单位: Yamagata University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12680587/
• 关键词: farnesyl diphosphate synthase; substrate analog; mutated enzyme; thermostable enzyme; geranyl diphosphate; dimethylallyl diphosphate; substrate specificity; isopentenyl diphosphate; ファルネシル二リン酸合成酵素; ゲラニル二リン酸; ジメチルアリル二リン酸; イソペンテニル二リン酸; ファルネシル二リン

Farnesyl diphosphate (FPP) synthase catalyze the condensation of isopentenyl diphosphate (IPP) with allylic primer to give FPP as final product. The FPS from pig liver has been successfully applied to syntheses of bioactive compounds. Molecular cloning and expression of the gene for thermostable FPS from Bacillus stearothermophilus made it possible to produce sufficient amounts of the enzyme. However it can hardly accept the substrate analogs having oxygen atom in their chain, which are easily accepted by the pig the liver enzyme. There may be some limitations in the thermostable enzyme for applying to organic synthesis. We constructed the FPP syntheses (FPSs) from Bacillus stearothermophilus, in which Tyr-81 was substituted with Ser (S), Arg (R), Asp (D), or Gly (G). Interestingly the substrate specificities of the mutated enzymes have been dramatically altered. They can easily accept the substrate analogs having oxygen atom. These results may suggest the application of the prenyltransferases to organic synthesis is more available.

Farnesyl二磷酸（FPP）合酶催化异戊烯基双磷酸（IPP）与烯丙基底漆的凝结，以将FPP作为最终产物。猪肝的FPS已成功应用于生物活性化合物的合成。来自芽孢杆菌的热稳定FPS基因的分子克隆和表达，使得可以产生足够量的酶。但是，它几乎无法接受其链中具有氧原子的底物类似物，猪很容易被肝酶接受。适用于有机合成的热稳定酶可能存在一些局限性。我们用芽孢杆菌构造了bacilothothothomophilus的FPP合成（FPS），其中Tyr-81用Ser（S），Arg（r），ASP（d）或Gly（G）代替Tyr-81。有趣的是，突变酶的底物特异性已经发生了巨大变化。它们可以轻松接受具有氧原子的底物类似物。这些结果可能表明将前转移酶应用于有机合成。

项目成果

M.Nagaki et al.,: "Artificial Substrates for Recombinant Undecaprenyl Diphosphate…"Journal of Holeuslar Catolysis B : Enzymatic. 9. 33-38 (2000)

M. Nagaki 等人，：“重组十一碳二烯基二磷酸的人工底物……”Holeuslar 催化 B 杂志：酶学。

M. Nagaki, S. Sato, Y. Maki, T. Nishino, and T. Koyama: "An Artificial substrate for undecaprenyl diphosphate synthase from Micrococcus luteus B-P 26"Catalysis Communication. Vol 1. 33-35 (2000)

M. Nagaki、S. Sato、Y. Maki、T. Nishino 和 T. Koyama：“来自藤黄微球菌 B-P 26 的十一异戊二烯基二磷酸合酶的人工底物”催化通讯。

M.Nagaki et al.: "Substrate specificityl of thermostable farnesy diphosphate synthase"J.Mol.Catelysis Bi Enzymatic. (in press). (2002)

M.Nagaki 等人：“热稳定性法呢基二磷酸合酶的底物特异性”J.Mol.Catethesis Bi Enzymatic。

M.Nagaki et al.: "An artificial substrate for undecaprenyl diphosphate synthase from M.Letus B-P 20"Catalysis Communication. 1. 33-35 (2000)

M.Nagaki 等人：“来自 M.Letus B-P 20 的十一异戊二烯基二磷酸合酶的人工底物”催化通讯。

M.Nagaki et al.: "An artificial substrate for undecaprenyl diphosphate synthase from M.Luteus B-P-20"Catalysis Communication. 1. 33-35 (2000)

M.Nagaki 等人：“来自 M.Luteus B-P-20 的十一异戊二烯基二磷酸合酶的人工底物”催化通讯。