CRITICAL ASSESSMENT OF THE PROTON INVENTORY METHOD

质子清单方法的严格评估

• 批准号: 2185016
• 负责人: ALEXANDER J KRESGE
• 金额: $ 9.16万
• 依托单位: UNIVERSITY OF TORONTO
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-05-01 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2185016
• 关键词: active sites; benzoates; chemical association; chemical kinetics; chymotrypsin; cyclodextrins; deuterium oxide; enzyme mechanism; enzyme model; enzyme substrate; enzyme substrate complex; hydrogen; hydrogen bond; hydrogen transport; hydrolysis; nonradiation isotope effect; nuclear magnetic resonance spectroscopy; protonation; site directed mutagenesis; solvents; ultraviolet spectrometry; water; water solution

This Application describes a program of research designed to provide a critical evaluation of the use of solvent isotope effects in H2O-D2O mixtures, the "proton-inventory" method, in studies of enzyme-catalyzed reactions. Such an assessment is necessary because serious questions have been raised about the ability of the proton-inventory method to enumerate catalytically active hydrogens in enzymic reactions, a purpose for which it has been used extensively and through which much of what is believed to be known about acid-base catalysis in enzymic systems is based. The proposed research consists of three parts: (1) examination of cyclodextrin-based models, (2) accurate determination of isotopic fractionation factors, and (3) studies with enzymes modified by site-directed mutagenesis. The results obtained would provide fundamental understanding of solvent isotope effects in H2O-D2O mixtures, and that would furnish practical guidelines for the correct application of the proton-inventory method to enzymic systems, and also supply criteria by which to judge the validity of conclusions reached through previous applications of the method.

该应用程序描述了一项研究计划，旨在提供 对H2O-D2O中溶剂同位素效应的使用的批判性评估 混合物，“质子构成”方法，用于酶催化的研究 反应。 这样的评估是必要的，因为严重的问题有 提出了关于质子 - 陈述方法枚举的能力的提高 酶促反应中的催化活性氢，其目的 已被广泛使用，并且被认为是的大部分 关于酶系统中酸碱催化的已知。 提议 研究由三个部分组成：（1）基于环糊精的检查 模型，（2）准确确定同位素分级因子和 （3）通过定向诱变修饰的酶的研究。 这 获得的结果将提供对溶剂同位素的基本理解 在H2O-D2O混合物中的影响，这将提供实用的指南 为了正确应用质子构成质子的方法 系统，还提供判断有效性的标准 结论通过该方法的先前应用得出。