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Synthesis of water-soluble selenium reagents and their applications to protein chemistry

水溶性硒试剂的合成及其在蛋白质化学中的应用

基本信息

批准号：
13640525
负责人：
IWAOKA Michio
金额：
$ 2.24万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13640525/
关键词：
selenium protein redox disulfide bond folding 酸化還元剤酸化還元試剤

项目摘要

For folding study of proteins with disulfide bonds (SS bonds), water soluble sulfur reagents are normally used. In this research project, new selenium compounds have been synthesized and applied to oxidative folding experiments of proteins. This study is based on a general chemical concept that selenium is more redox-acitive than sulfur.Two-types of water-soluble selenium reagents were synthesized. When one of the reagents was used for folding experiments, the reagent reacted with a reduced form of bovine pancreatic ribonuclease A rapidly (within 10 sec) and quantitatively to form protein SS bonds. To investigate details of the rapid processes, a quench-flow equipment was constructed. The kinetic experiments using the quench-flow equipment revealed that the reaction between the protein and the selenium reagent completes within 1 sec and also that the SS formation proceeds during 50 to 100 msec. The results suggested that the selenium reagent may be able to trap folding intermediates that are formed rapidly in the beginning of folding and are alive in a short period. Thus, the selenium reagents would be useful for oxidative folding study of proteins.In this research project, database analysis of weak interactions in PDB and ab initio molecular orbital calculations of single amino acids have been also carried out. S…O interactions were discovered in protein structures. Conformational preferences of a single amino acid in water were suggested to be important in the folding of a polypeptide chain to secondary structures.

对于具有二硫键（SS键）的蛋白质的折叠研究，通常使用水溶性硫试剂。本课题合成了新的硒化合物，并将其应用于蛋白质的氧化折叠实验。这项研究是基于一个普遍的化学概念，即硒比硫具有更强的氧化活性。合成了两种水溶性硒试剂。当其中一种试剂用于折叠实验时，该试剂与还原形式的牛胰腺核糖核酸酶a快速（在10秒内）定量反应，形成蛋白质SS键。为了研究快速过程的细节，构建了一个淬火流设备。用急流装置进行的动力学实验表明，蛋白质与硒试剂的反应在1秒内完成，SS的形成在50 ~ 100秒内完成。结果表明，硒试剂可能能够捕获在折叠开始时迅速形成并在短时间内存活的折叠中间体。因此，硒试剂可用于蛋白质氧化折叠的研究。本课题还进行了PDB中弱相互作用的数据库分析和单氨基酸从头算分子轨道计算。在蛋白质结构中发现了S…O相互作用。单个氨基酸在水中的构象偏好被认为是多肽链折叠成二级结构的重要因素。