New Probe for Transient Substrate Radicals in Enzymic Catalysis

酶催化中瞬态底物自由基的新探针

• 批准号: 9118911
• 负责人: Bruce Branchaud
• 金额: $ 2万
• 依托单位: University of Oregon Eugene
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-04-01 至 1993-09-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9118911&HistoricalAwards=false
• 关键词: New; Probe; Transient; Substrate; Radicals

In recent years evidence has been accumulating that many enzymes proceed through radical mechanisms. Most reactions involve redox cofactors for which radical intermediates are plausible based on model studies of cofactor chemistry. New types of mechanisms have been proposed recently involving radicals on amino acid side chains. Although there has been much compelling indirect evidence consistent with H-atom abstraction from substrates by protein-derived radicals, to date there has been no direct evidence. Galactose oxidase from the fungus Dactylium dendroides catalyzes the oxidation of primary alcohols with O2, producing aldehydes and H2O2. Based on recent EPR, resonance Raman, and X-ray crystallographic structural information and on extensive preliminary studies with mechanism-based inactivating substrate analogs, a new mechanism for galactose oxidase catalysis is proposed featuring rate-determining H-atom abstraction by a tyrosine radical from the alcohol CH2. The reaction mechanism of galactose oxidase catalysis will be studied in the proposal research. The resulting detailed reaction mechanism should become a paradigm for substrate activation using protein-centered radicals for H- atom abstraction from substrates. Such fundamental information will be useful in understanding more biomedically relevant enzymes, such as ribonucleotide reductases, which are more difficult to study due to stringent substrate specificities.

近年来，越来越多的证据表明 酶通过自由基机制进行。 大多数反应 涉及氧化还原辅因子，对于氧化还原辅因子， 基于辅因子化学的模型研究似乎是合理的。 新 最近提出了几种机制， 氨基酸侧链上的自由基。 虽然出现 许多令人信服的间接证据与氢原子 通过蛋白质衍生的自由基从底物中提取， 至今没有直接证据。 半乳糖氧化酶 真菌Dactyelodendroides催化氧化 伯醇与O2反应，生成醛和H2 O2。 基于 在最近的EPR，共振拉曼，和X射线晶体 结构信息和广泛的初步研究 通过基于机制的灭活底物类似物，一种新的 半乳糖氧化酶的催化机制提出了特点 酪氨酸自由基对氢原子的夺去速率决定作用 醇CH 2。 半乳糖氧化酶的反应机理 催化将在提案研究中进行研究。 的 由此产生的详细反应机制应该成为一个范例 对于底物活化，使用蛋白质为中心的自由基进行H- 从基质中提取原子。 这些基本信息 将有助于理解更多与生物医学相关的 酶，如核糖核苷酸还原酶，这是更多的 由于严格的底物特异性而难以研究。