MAGNETIC AND MODELING STUDIES OF CYTOCHROME OXIDASE

这项提案分为两部分，延续了我们之前的工作。第一部分 涉及Faraday和 全氧化(休眠)各种衍生物的SQUID方法学 细胞色素C氧化酶的性质和性质的测定 铁磁或反铁磁交换的大小(正/负J值) 铁(III)和铜(II)在双核上的相互作用 (Cyt.a3-3(桥)Cuu2)活性部位。这两种哺乳动物的氧化酶 (牛)和细菌(T.thermophilus)来源将被研究。新的 建议我们继续研究的酶的衍生物是 休息(厌氧准备)、休息.F-、休息.H202和“脉冲” 牛氧化物酶的衍生物及其休眠和休眠.CN-形式 嗜热链霉菌的酶。这些额外的磁化学结果 衍生品，以及我们之前对RESTING、RESTING、CN-、 和休息。牛氧化酶的HCO2-衍生物将提供进一步的洞察力 成为催化参与的、桥接的活跃分子的领先候选人 站点结构：(1)Mu-oxo病例，(Cyt.a3-3-0-Cuu2)，(2)a Mu-Mercapto病例(Cyt.a3-3-S-Cuu2)，程度较轻的 Mu-咪唑拉多型(Cyt.a3-3(IMiD)Cuu2)。第二部分将涉及 合成、结构表征(X-射线结晶学和EXAFS) 和研究(磁化学、RR、EPR和穆斯堡尔谱) 各种[(TPP)FeIII(桥)CuII]模型化合物(或有用的合成化合物 类似物)，其中桥是氧化酶的可能性：氧代，硫醇， 咪唑、CN-、HCO2-、F-和H202。通过初步的和持续的 工作中，已有20种新的异质双金属物种问世 并产生与酶活性位点有关的信息 结构以及由此产生02 4H 4E-的机制 2H20酶反应被催化。

This proposal is in two parts and continues our previous work. Part I involves full-temperature (4.2-300K) magnetochemical studies by Faraday and SQUID methodologies of various derivatives of fully-oxidized (resting) cytochrome c oxidase for the purpose of determining the nature and magnitude (plus/minus J value) of the ferro- or antiferromagnetic exchange interaction between (Fe(III) and Cu(II) at the binuclear (Cyt.a3-3+(bridge)Cuu2+) active site. Oxidases from both mammalian (bovine) and bacterial (T. thermophilus) sources will be studied. The new derivatives of the enzyme proposed for our continuing studies are the resting (anaerobically prepared), resting.F-, resting.H202 and "pulsed" derivatives of bovine oxidase and the resting and resting.CN- forms of the T. thermophilus enzyme. Magnetochemical results for these additional derivatives, along with our previous findings for the resting, resting.CN-, and resting.HCO2- derivatives of bovine oxidase will lend further insight into the leading candidates for the catalytically-involved, bridged active site structure: (1) a Mu-oxo case with (Cyt.a3-3+-0-Cuu2+), (2) a Mu-mercapto case with (Cyt.a3-3+-S-Cuu2+), and to a lesser extent, a Mu-imidazolato case with (Cyt.a3-3+(imid)Cuu2+). Part II will involve the synthesis, structural characterization (x-ray crystallography and EXAFS) and study (magnetochemistry, RR, epr, and Mossbauer spectroscopy) of various [(TPP)FeIII(bridge)CuII] model compounds (or useful synthetic analogues) where the bridges are the oxidase possibilities: oxo, mercapto, imidazolato, CN-, HCO2-, F-, and H202. Through preliminary and on-going work, twenty of these novel heterobimetallic species are already in hand and are yielding information pertaining to the oxidase active site structure and thereby the mechanism whereby the 02 + 4H+ + 4e- produces 2H20 enzyme reaction is catalyzed.