Transition State Analysis of Enzymatic Reactions

酶促反应的过渡态分析

• 批准号: 7268940
• 负责人: Vern L. Schramm
• 金额: $ 49.93万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7268940
• 关键词: 5-carboxy-methylaminomethyl-2' -O-methyluridine; Bees; Binding; Biochemical Reaction; Calcium Binding; Cell physiology; Complex; Crystallography; Development; Disease; Electrons; Enzymes; Generations; Goals; Investigation; Isoenzymes; Isotopes; Kinetics; Lasers; Mammals; Measurement; Mus; Nature; Nucleoside Hydrolases; Object Attachment; Parasites; Pattern Recognition; Proteins; Purine-Nucleoside Phosphorylase; Purines; Research Project Grants; T-Lymphocyte; Testing; Training; Water; analog; clinically relevant; design; enzyme structure; inhibitor/antagonist; interest; novel; purine; theories

The goals of this research project are to characterize the transition states of biologically significant enzymes of N-ribosyl bond scission. The transition states are used as atomic templates to develop the theory and practice of transition state inhibitor design. Transition state information and transition state analogies are used to investigate the nature of enzyme-bound transition states. The enzymes of interest include the protozoan nucleoside N-ribohydrolase isozymes and purine nucleoside phosphorylase. Protozoan parasites are purine auxotrophs and use three isozyme of N-ribohydrolases for purine salvage. The enzymes are not found in mammals. Purine nucleoside phosphorylase is widely distributed and is essential for normal T-cell function. Transition state structures for these enzymes have bee determined by kinetic isotope effect measurements using normal mode semiemperical and ab initio structural analysis. The first generation of transition state inhibitors have been synthesized and will be characterized. Experimental approaches will include: 1) laser and infrared spectroscopic investigation of the bound calcium-water center in the nucleoside hydrolases; NMR and Raman spectra of bound transition state inhibitors using isotope-edite difference analysis; and free-electron induced infrared lasers to study the actual enzymatic transition state; 2) x-ray crystallography of complexes with substrate and transition state analogues to define changes in protein structur in empty enzyme, Michaelis and transition state complexes; 3) development of theory for predicting binding energies of transition state inhibitors using trained pattern recognition; synthesis and testing of likely transition state inhibitors. The most powerful inhibitors will be tested for function in mice. The results of these studies are expected to give novel information about the nature of enzymatic transition states, provide new theory for the design of transition state inhibitors, and to provide powerful transition state inhibitors for several clinically relevant disorders.

该研究项目的目标是表征 N-核糖基键的生物学意义酶的过渡态 分裂。 过渡态被用作原子模板来开发 过渡状态抑制剂设计的理论和实践。 过渡状态 信息和过渡状态类比用于研究 酶结合过渡状态的性质。 感兴趣的酶包括 原生动物核苷N-核糖酶同工酶和嘌呤核苷 磷酸化酶。 原生动物的寄生虫是嘌呤的合子，使用三个 N-核糖液酶的同工酶用于嘌呤挽救。 找不到酶 在哺乳动物中。 嘌呤核苷磷酸化酶被广泛分布，为 对于正常的T细胞函数必不可少的。 这些的过渡状态结构 酶已通过动力学同位素效应测量来确定蜜蜂 正常模式的半经典和从头开始结构分析。 第一个 过渡状态抑制剂的产生已经合成，将是 特征。 实验方法将包括：1）激光和红外线 对结合钙 - 水中心的光谱研究 核苷水解酶； NMR和拉曼频谱的结合过渡状态 使用同位素差异分析的抑制剂；和自由电子 诱导红外激光研究实际酶促过渡态； 2） 具有底物和过渡状态的复合物的X射线晶体学 定义空酶中蛋白质结构变化的类似物，迈克尔斯 和过渡状态综合体； 3）预测理论的发展 使用训练的模式的过渡状态抑制剂的结合能 认出;可能过渡状态抑制剂的合成和测试。 最强大的抑制剂将在小鼠中测试功能。 这 这些研究的结果有望提供有关有关的新信息 酶过渡状态的性质，为设计提供了新的理论 过渡状态抑制剂，并提供强大的过渡状态 几种临床相关疾病的抑制剂。

项目成果

Transition state analogues in quorum sensing and SAM recycling.

• DOI: 10.1093/nass/nrn038
• 发表时间: 2008-01-01
• 期刊: Nucleic acids symposium series (2004)
• 作者: Schramm, Vern L;Gutierrez, Jemy A;Furneaux, Richard H
• 通讯作者: Furneaux, Richard H

Immucillins in custom catalytic-site cavities.

定制催化位点腔中的免疫西林。

• DOI: 10.1016/j.bmcl.2008.08.047
• 发表时间: 2008
• 期刊: Bioorganic & medicinal chemistry letters
• 影响因子: 2.7
• 作者: Murkin,AndrewS;Clinch,Keith;Mason,JenniferM;Tyler,PeterC;Schramm,VernL
• 通讯作者: Schramm,VernL