Origine and Molecular Evolutionary Engineering of the Enzyme-Coenzyme Systems

酶-辅酶系统的起源和分子进化工程

• 批准号: 08680683
• 负责人: YOSHIMURA Tohru
• 金额: $ 1.54万
• 依托单位: KYOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08680683/
• 关键词: coenzyme; molecular evolution; pyridoxal phosphate; stereochemistry; aminotransafearse; lyase; glutamate racemase

The stereospecificities for the hydrogen transfer between C-4' of the cofactor and substrate in the transamination catalyzed by various pyridoxal 5'-phosphate (PLP)-dependent enzymes have been studied. The stereospecificities reflect the active-site structures of the enzymes, especially the topographical situation of a coenzyme-substrate Schiff base and a catalytic base for the hydrogen transfer. We found that three types of enzymes catalyzing the reaction stereospecifically on either si-or re-face of the planar intermediate, and alternatively non-stereospecifically on both faces. The classification of PLP-enzymes based on the stereospecificcity for the hydrogen transfer coincident with that based on their primary and three dimensional structures. These suggest that PLP-enzymes have been evolved from at least three different ancestral proteins. These studies lead us the speculation that enzymes requiring coenzymes were created by the combination of the inherently active coenzymes and protein offering the place of the reactions. We attempted to produce a new enzyme by combining a hemin and glutamate recemase showing a significant similarity with mammalian myoglobins in their primary structures.

研究了5 ′-磷酸吡哆醛（PLP）依赖性酶催化转氨反应中辅因子C-4 ′与底物间氢转移的立体专一性。立体专一性反映了酶的活性中心结构，特别是辅酶-底物席夫碱和氢转移催化碱的拓扑位置。我们发现三种类型的酶催化反应的平面中间体的任何一个表面或背面的立体专一性，或者非立体专一性的两面。基于氢转移立体专一性的PLP-酶分类与基于其一级结构和三维结构的分类是一致的。这些表明PLP酶已经从至少三种不同的祖先蛋白进化而来。这些研究使我们推测，需要辅酶的酶是由固有活性的辅酶和提供反应场所的蛋白质结合而产生的。我们试图通过结合氯化血红素和谷氨酸再合成酶来产生一种新的酶，这种酶在一级结构上与哺乳动物肌红蛋白具有显著的相似性。

项目成果

吉村,徹: "D-アミノ酸の役割 生理作用と代謝をめぐって" 現代化学. 12号. 14-19 (1996)

吉村彻：“D-氨基酸的作用：生理效应和代谢”Gendai Kagaku No. 12. 14-19 (1996)。

Yoshimura, T.et al.: "Stereospecificity for the Hydrogen Transfer and Molecular Evolution of Pyridoxal Enzymes" Biosci.Biotech.Biochem.60 (2). 181-187 (1996)

Yoshimura, T.等人：“吡哆醛酶的氢转移和分子进化的立体特异性”Biosci.Biotech.Biochem.60 (2)。

Jhee,K.‐H.: "Stereospecificity of Thermostable Ornithine 5‐Aminotransferase for the Hydrogen Transfer in the L‐ and D‐Ornithine Transamination" Biochemistry. 35巻30号. 9792‐9796 (1996)

Jhee, K.-H.：“L-和 D-鸟氨酸转氨作用中氢转移的热稳定鸟氨酸 5-氨基转移酶的立体特异性”，《生物化学》，第 35 卷，第 30 期。9792-9796 (1996)

Yoshimura, T.: "Structure and Function of Bacterial D-Amino Acid-Related Enzymes Enzymes" Nippon Nougeikagaku kaishi. 70 (1). 15-20 (1996)

Yoshimura, T.：“细菌 D-氨基酸相关酶的结构和功能”Nippon Nougeikagaku kaishi。

Liu, L.et al.: "Cloning and Expression of the Glutamate Racemase Gene of Bacillus pumilus" J.Biochem.121 (6). 1155-1161 (1997)

Liu, L.等人：“短小芽孢杆菌谷氨酸消旋酶基因的克隆和表达”J.Biochem.121 (6)。