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Structural Regulation Mechanism for Reactivity in Metalloproteins

金属蛋白反应性的结构调控机制

基本信息

批准号：
07309006
负责人：
MORISHIMA Isao
金额：
$ 7.23万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07309006/
关键词：
Peroxidase Ferredoxin Nitrite Reductase Oxgen Snsor Fix L Cytochrome c Oxidase Cytochrome P450 酸素センサーFixL チトクロム酸化酵素

项目摘要

The Primary results in this research project are as follows :1)The extensive mutation in horseradish peroxidase has revealed that the highly conserved hydrogen network in the distal site plays a key role in catalytic activity in peroxidases. The detailed molecular mechanism in peroxidase reaction was discussed.2)The crystal structure of ARP,which shows specific high reactivity for luminol, was resolved and the substrate binding site was proposed. By using time-resolved resonance Raman spectroscopy, we examined the intermediate species in the reaction ARP and compared them with those of horseradish peroxidase.3)We followed the reaction of cytochrome c oxidase with hydrogen peroxide and idetified some key inter mediates in the reaction.4)The Regulation mechanism for the terminal oxidase in Escherichia coli was investigated by using the mutants.5)The electron transfer in nitrite reductase, cytochrome cd_1, was examined by pulse radiolysis and reduction mechanism for the enzyme was proposed.6)The systematic mutation in ferredoxin showed the specific interaction sites between ferredoxin and sulfite reductase.7)We resolved crystal structure of another nitrite reductase, P450nor, and discussed the molecular mechanism for reduction of nitrite by the enzyme.8)EPR was utilized for the investigation of the interaction between cytochrome P450cam and its electron donor, putidaredoxin. Some specific interactions were elucidated.

本研究的主要结果如下：1）辣根过氧化物酶的广泛突变揭示了其末端高度保守的氢网络在过氧化物酶的催化活性中起着关键作用。2）解析了对鲁米诺具有特异性高反应活性的阿普的晶体结构，提出了ARP的底物结合位点。利用时间分辨共振拉曼光谱研究了阿普的中间产物，并与辣根过氧化物酶的中间产物进行了比较; 3）跟踪了细胞色素c氧化酶与过氧化氢的反应，确定了反应中的关键中间产物; 4）利用突变体研究了大肠杆菌末端氧化酶的调控机制; 5）亚硝酸还原酶的电子传递，6）铁氧还蛋白的系统突变揭示了铁氧还蛋白与亚硫酸盐还原酶的特异性相互作用位点; 7）我们解析了另一种亚硝酸盐还原酶P450 nor的晶体结构，探讨了该酶还原亚硝酸盐的分子机理。用电子顺磁共振（EPR）研究了细胞色素P450 cam与其电子供体putidaredoxin的相互作用。阐明了一些特定的相互作用。