Structures & Redox Chemistry in Sulfinic Acid Reduction

描述（由申请人提供）：亚硫酸还原酶，sulfiredoxin (Srx)，最近被确定为酵母中负责还原几种氧化失活的过氧化物还毒素（Prxs）中亚硫酸部分（Cys-SO2-）的酶。这一发现打破了Prx酶家族过度氧化不可逆性的教条。此外，过度氧化Prxs的ATP和Mg2+依赖性修复或“逆转录”可能调节这些酶作为过氧化氢介导的细胞内信号传导的调节剂的作用。基于对多种人类Prx在体内氧化状态的分析，Srx对不同Prx亚型的修复似乎以不同的速率进行，尽管它们的整体序列高度一致。我们最近对人类Srx与磷酸或ADP复合物的结构测定揭示了一个新的蛋白折叠和一个新的核苷酸结合基序。其他初步数据已经证实了酵母Srx作用方案的某些方面，尽管我们的结果中有几个令人惊讶的地方（分离出一个二硫键结合的中间体而不是硫亚硫酸盐连接的中间体，以及谷胱甘肽而不是硫氧还蛋白还原Srx:Prx复合物）表明需要进一步的研究。

DESCRIPTION (provided by applicant): The sulfinic acid reductase, sulfiredoxin (Srx), was recently identified as the enzyme in yeast responsible for the reduction of the sulfinic acid moiety (Cys-SO2-) within several oxidatively-inactivated peroxiredoxins (Prxs). This discovery shattered the dogma of the irreversibility of overoxidation for the Prx enzyme family. Moreover, the ATP- and Mg2+-dependent repair or "retroreduction" of the overoxidized Prxs may modulate the role of these enzymes as regulators of hydrogen peroxide-mediated intracellular signaling. Based on the analysis of the in vivo oxidation state for a variety of human Prxs, repair of different Prx isoforms by Srx appears to proceed at different rates, in spite of their high overall degree of sequence identity. Our recent structure determinations of human Srx in complex with either phosphate or ADP have revealed a new protein fold and a novel nucleotide binding motif. Other preliminary data have confirmed some aspects of the proposed scheme for yeast Srx action, although several surprises in our results (the isolation of a disulfide-bonded rather than thiosulfinate-linked intermediate, and the reduction of the Srx:Prx complex by glutathione rather than thioredoxin) have suggested that further investigation is required. The goals of this proposal are: to determine the crystal structures of human Srx in complex with cofactors and human Prxs (Aim 1); and to carry out steady-state and partial turnover experiments coupled with site-directed mutagenesis to elucidate the nature and reaction rates of intermediates, and to characterize residues critical to catalysis (Aims 2 & 3). These investigations will contribute significantly to our understanding of the molecular origins of sulfinic acid reductase action and the novel sulfur chemistry involved in this process.