High Resolution X-ray Study of Cofactor Dependent Enzyme and Biocatalyst Design

利用x射线晶体学方法测定了谷氨酰胺氨基转移酶、变构CTP合成酶、plp依赖性酶和fad依赖性酶等天然复合催化剂的高分辨率结构。分析了底物识别、诱导配合和反应机制，以了解使这些酶作为生物催化剂的分子设计。天然生物催化剂的三维结构和活性位点设计。(1)咪唑甘油磷酸合成酶。通过连接谷氨酰胺酶和合成酶位点的分子通道进行氨转移的机制已被提出。(2)CTP（胞苷5′-三磷酸）合成酶。研究了ATP和utp依赖性构象变化诱导的氨隧穿、变构GTP激活和酶激活的机制。(3) Argininosuccinate合成酶。为了进一步阐明该反应的化学性质，进行了底物识别机理和伪时间分辨x射线分析。(4)醌血红蛋白胺脱氢酶。一种新的奎宁辅因子和极不寻常的硫醚交叉桥已被确定。plp依赖性酶的三维结构。(1) Histidinol-phosphate转氨酶。分析了双底物识别的活性位点设计。(2)支链氨基酸转氨酶。研究了双底物识别及其反应机理。(3)苏氨酸合成酶。预期催化的反应将通过plp依赖性酶所利用的所有中间体进行。提出了该反应的立体化学性质。(4)谷氨酰胺/ kunerenine转氨酶。在哺乳动物中，这种酶参与中枢神经系统的活动。研究了底物识别和诱导拟合现象。fad依赖性酶的三维结构。(1)中链酰基coa氧化酶。在与同系物（酰基辅酶a脱氢酶）结构比较的基础上，分析了该氧化酶的分子设计。(2)中链酰基辅酶a脱氢酶作为过渡态类似物。根据配合物的x射线结构和MO计算，研究了反应的过渡态。少

High-resolution Structures of natural composite catalysts such as glutamine amidotransferase, and allosteric CTP synthetase, PLP-dependent enzymes, and FAD-dependent enzymes have been determined by X-ray crystallographic methods. The substrate recognition, induced fit, and reaction mechanism have been analyzed to understand the molecular design which enables these enzymes to act as biocatalysts.1. Three-dimensional structures of natural biocatalysts and active-site designs. (1)Imidazole glycerol phosphate synthetase. The ammonia transfer mechanism through a molecular channel linking glutaminase and synthase sites has been proposed. (2)CTP (cytidine 5'-triphosphate) synthetase. The mechanism for ammonia tunneling, allosteric GTP activation, and enzyme activation induced by ATP and UTP-dependent conformational change has been examined. (3)Argininosuccinate synthetase. The substrate recognition mechanism and pseudo time-resolved X-ray analysis has been performed in order to clarify the st … More ereochemistry of the reaction. (4) Quinohemoprotein aminedehydrogenase. A novel quinine cofactor and highly unusual thioether cross-bridges have been identified.2. Three-dimensional structures of PLP-dependent enzymes. (1)Histidinol-phosphate aminotransferase. The active-site design for double substrate recognition has been analyzed. (2)Branched-chain amino acid aminotransferase. The double substrate recognition and reaction mechanism have been elucidated. (3)Threonine synthase. The reaction catalyzed is expected to proceed through all the intermediates utilized by PLP-dependent enzymes. Stereochemistry of the reaction has been proposed. (4)Glutamine/kunerenine aminotransferase. In mammals, the enzyme is involved in CNS events. The substrate recognition and induced fit phenomena have been examined.3. Three dimensional structures of FAD-dependent enzymes. (1)Medium-chain acyl-CoA oxidase. On the basis of structural comparison with the homologue (acyl-coA dehydrogenase), the molecular design of the oxidase has been analyzed. (2)Medium-chain acyl-CoA dehydrogenase as the transition state analogue. The transition state of the reaction has been examined based on the X-ray structure and MO calculation of the complex. Less