SINGLE PROTON EXCHANGE KINETICS IN PROTEINS

蛋白质中的单质子交换动力学

• 批准号: 3273760
• 负责人: CLARE K WOODWARD
• 金额: $ 15.29万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1979
• 资助国家: 美国
• 起止时间: 1979-04-01 至 1990-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3273760
• 关键词: acidity /alkalinity; chemical reaction; conformation; crosslink; deuterium; hydrogen transport; isomorphous substitution; molecular rearrangement; nuclear magnetic resonance spectroscopy; peptide chemical synthesis; stop flow technique; thermodynamics; trypsin inhibitors

The general goal of this project is to determine the internal motions of proteins by which atoms buried in the matrix of folded proteins are accessible to solvent. This is measured by the hydrogen isotope exchange kinetics of solvent hydrogen atoms with labile protein protons, most of which are peptide amide protons. The hydrogen isotope exchange kinetics of peptide NN's in folded proteins are distributed over 6-10 orders of magnitude. The most rapidly exchangeing NH's are on the surface, and the slowest exchanging NH's tend to be in buried sections of Beta -sheet. Although buried NH's exchange slower than model compounds, their finite exchange rates demonstrate that interior atoms in native proteins have some probability of being exposed to solvent. This immediately implies that the protein fluctuates to render tightly packed, buried atoms accessible to solvent. Recently our laboratory has made the exciting findings that the exchange rates of NH's on the surface of bovine pancreatic trypsin inhibitor (BPTI) vary from 3-fold to 2000-fold slower than model compounds, that their pHmin values vary over greater than 2 pH units, and that some have pHmin of less than 1. It has been commonly assumed that surface NH atoms (not H-bonded and with finite static accessibility) exchange with rates comparable to those in model peptides. The deviation in exchange rates and pHmin from model compounds then taken on special interest as a reflection of the dynamic structure of the protein-solvent interface. The specific aims of this research proposal are 1) to characterize the hydrogen exchange kinetics of NH's in BPTI with intermediate exchange rates, 2) to measure the effect of trypsinogen and trypsinogen/Ile-Val binding on the exchange rates of BPTI NH's 3) to compare the exchange rate of individual BPTI NH's in solution with their exchange rates in the crystalline and powder forms, and 4) to measure the exchange kinetics of water buried in the BPTI-trypsin complex with solvent water.

这个项目的总体目标是确定的内部运动， 蛋白质，其中原子埋在折叠的蛋白质基质中， 易溶于溶剂。 这是通过氢同位素交换来测量的 溶剂氢原子与不稳定蛋白质质子的动力学， 它们是肽酰胺质子。 氢同位素交换动力学 折叠蛋白质中的肽NN分布在6-10个数量级上， 大小 最快交换的NH在表面， 最慢NH交换倾向于在β折叠的埋藏部分。 虽然埋藏NH的交换比模型化合物慢，但它们的有限 交换率表明，天然蛋白质中的内部原子具有一些 暴露于溶剂的可能性。 这直接意味着， 蛋白质波动，使紧密包装，埋藏的原子， 溶剂后 最近，我们的实验室取得了令人兴奋的发现， 牛胰胰蛋白酶抑制剂（BPTI）表面NH的速率 比模型化合物慢3倍至2000倍， 值在大于2个pH单位的范围内变化，并且有些具有小于 超过1. 通常假设表面NH原子（非H键合的 和有限的静态可达性），汇率与 那些在模型肽中。 汇率和pH最小值与 模型化合物，然后采取了特别的兴趣，作为反映的 蛋白质-溶剂界面的动态结构。 本研究建议的具体目标是：1）描述 中间交换BPTI中NH ′ s氢交换动力学 率，2）测量胰蛋白酶原和胰蛋白酶原/Ile-Val的作用 对BPTI NH的汇率有约束力3）比较汇率 的个别BPTI NH的解决方案，其汇率在 晶体和粉末形式，和4）测量交换动力学， 用溶剂水包埋在BPTI-胰蛋白酶复合物中的水。