Elucidation of recognition sites of lipase toward its development as chemical catalysis

阐明脂肪酶的识别位点以促进其化学催化的发展

• 批准号: 10650859
• 负责人: KOMETANI Tadashi
• 金额: $ 1.28万
• 依托单位: Toyama National College of Technology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10650859/
• 关键词: lipase; hydrolytic activity; enantioselectivity; Candida rugosa; 立体選択性; カンジダ ルゴサ; カンジタルゴサ

The aim of this project is to realize and control an enzymatic reaction as a chemical reaction. The research results are as follows ;(1) Candida rugosa lipase (CRL) was treated with several concentrations of 2-propanol in water and the changes of the amount of the recovered protein, the hydrolytic activity, and the enentioselectivity toward methyl 2-phenylpropanoate were studied. The results shows that the conformational change of CRL occurred in 30% 2-propanol treatment.(2) The sizes of the large, medium, and small substituent recognition sites (L, M, and S pocket, respectively) in CRL were roughly estimated by measuring the specific hydrolytic activity against several p-nitrophenyl esters. These relative sizes were assessed as L pocket > phenyl, ethyl > M pocket > methyl > S pocket > hydrogen.(3) The highly enantioselective transesterification of 2-phenyl-1-propanol, which has not been efficiently resolved by lipase-catalyzed reactions, was attained by using vinyl esters having aromatic ring in acyl moiety as vinyl 3-phenylpropanoate.

本项目的目的是实现和控制作为化学反应的酶反应。主要研究结果如下：（1）研究了不同浓度的异丙醇对皱纹假丝酵母脂肪酶（Candida rugosa lipase，CRL）的蛋白质回收量、水解活性以及对2-苯基丙酸甲酯的对映选择性的影响。结果表明，30%异丙醇处理后CRL的构象发生了变化。(2)通过测量对几种对硝基苯基酯的比水解活性，粗略估计了CRL中大、中、小取代基识别位点（分别为L、M和S口袋）的大小。这些相对大小被评估为L口袋>苯基，乙基> M口袋>甲基> S口袋>氢。(3)以酰基中含有芳环的乙烯基酯作为3-苯基丙酸乙烯酯，实现了2-苯基-1-丙醇的高对映选择性酯交换反应，这一反应在脂肪酶催化下尚未得到有效解决。

项目成果

Michimasa Goto 他: "Biding site of Acyl Moiety in Ester Hydrolysis by Candida rugosa lipase"Journal of Molecular Catalysis B : Enzymatic. (in press).

Michimasa Goto 等人：“Candida rugosa 脂肪酶酯水解中酰基部分的结合位点”《分子催化杂志 B》：酶学（正在出版）。

Michimasa Goto, Masashi Kawasaki, Tadashi Kometani: "Enzymatic resolution of 2-phenyl-1-propanol by enantioselective hydrolysis of its ester having a bulky group in an acyl moiety"Journal of Molecular Catalysis B : Enzymatic. (in press).

Michimasa Goto、Masashi Kawasaki、Tadashi Kometani：“通过对映选择性水解其在酰基部分中具有大基团的酯来酶促拆分 2-苯基-1-丙醇”《分子催化杂志 B》：酶促。

Michimasa Goto 他: "Enzymatic Resolution of 2-Phenyl-1-propanol by Enantioselective Hydrolysis of Its Ester Having a Bulky Group in an Acyl Moiety"Journal of Molecular Catalysis B: Enzymatic. (in press).

Michimasa Goto 等人：“通过对酰基部分中具有大体积基团的酯进行对映选择性水解来酶促拆分 2-苯基-1-丙醇”，《分子催化杂志 B：酶促》（出版中）。

Masashi Kawasaki 他: "Lipase-Catalysed Transesterification of 2-Phenyl-1-Propanol with Vinyl Esters Having Aramatic Ring in Acyl Moiety"Tetrahedron Letters. 40・28. 5223-5226 (1999)

Masashi Kawasaki 等人：“脂肪酶催化的 2-苯基-1-丙醇与酰基部分具有芳香族环的乙烯基酯的酯交换”四面体快报 40・28（1999）。

Michimasa Goto 他: "Binding Site of Acyl Moiety in Ester Hydrolysis by Candida rugosa lipase"Journal of Molecular Catalysis B: Enzymatic. (in press).

Michimasa Goto 等人：“Candida rugosa 脂肪酶酯水解中酰基部分的结合位点”，分子催化杂志 B：酶学（出版中）。