RUI: Characterization of the Manganese Centers of Oxalate Oxidase from Ceriporiopsis subvermispora

本项目旨在阐明蛋白质环境如何调节重组草酸氧化酶中草酸降解的化学过程。草酸氧化酶催化草酸碳-碳键裂解生成二氧化碳和过氧化氢。虽然目前还没有关于C. subvermispora草酸氧化酶（CsOxOx）的结构信息，但序列数据和同源性模型表明，它是第一个被鉴定的催化该反应的含锰bicupin酶。最具特征的草酸氧化酶来自大麦和小麦。这些酶，也被称为germins，含有单个杯状结构域，因此被归类为单杯状结构域。有趣的是，CsOxOx与bicupin微生物草酸脱羧酶的序列同源性最大。本研究的短期目标是：1)表征草酸氧化酶对锰的依赖性；2)确定该酶的活性位点。这项研究有望有助于理解微妙的结构变化如何影响进化相关蛋白质的显着功能变化。表征CsOxOx的锰依赖性是重要的，以便将该酶置于其他草酸降解酶和其他cupin蛋白的背景下。确定CsOxOx的活性位点是一个重要的问题。如果只有一个锰中心介导催化，则提出了关于第二个锰结合结构域的功能（如果有的话）以及每个结构域的局部蛋白质结构导致差异反应性的程度的关键问题。更广泛的影响这个项目正在将盖恩斯维尔州立学院（佐治亚大学系统的两年制单位）从一个提供优秀科学课程的机构转变为一个也为学生提供参与有意义和重要的实验室研究的机会的机构。本科生在盖恩斯维尔州立学院（GSC）开展大部分工作，但也有机会与佛罗里达大学和佛罗里达州塔拉哈西的国家高磁场实验室的合作者合作。学生研究人员有望通过各种媒体向广大受众交流他们的研究经验。学生研究人员也有望在专业会议和地区教育界发表他们的研究结果和报告。本项目的资金用于购买、操作和维护GSC无法获得的研究设备。

This project seeks to elucidate how protein environment modulates the chemistry of oxalate degradation in recombinant oxalate oxidase from Ceriporiopsis subvermispora. Oxalate oxidase catalyzes the carbon-carbon bond cleavage of oxalate to yield carbon dioxide and hydrogen peroxide. Although there is currently no structural information available for oxalate oxidase from C. subvermispora (CsOxOx), sequence data and homology modeling indicate that it is the first manganese-containing bicupin enzyme identified that catalyzes this reaction. The best characterized oxalate oxidases are from barley and wheat. These enzymes, also known as germins, contain a single cupin domain and are therefore classified as monocupins. Interestingly, CsOxOx shares greatest sequence homology with the bicupin microbial oxalate decarboxylases. The short term goals of this research are to 1) to characterize the manganese-dependence of oxalate oxidase from C. subvermispora and 2) to identify the active site of the enzyme. This research is expected to contribute to the understanding of how subtle structural changes effect remarkable functional variation in evolutionarily related proteins. Characterizing the manganese-dependence of CsOxOx is significant in order to place this enzyme in the context of other oxalate degrading enzymes and that of other cupin proteins. Identifying the active site of CsOxOx is an important problem. If only one of the manganese centers mediates catalysis, critical questions are raised concerning the function (if any) of the second Mn-binding domain and the extent to which local protein structure in each domain results in differential reactivity.Broader ImpactsThis project is transforming Gainesville State College (a two-year unit of the University System of Georgia) from an institution that provides excellent coursework in the sciences into one that also provides students the opportunity to participate in meaningful and important laboratory research. Undergraduate students carry out most of this work at Gainesville State College (GSC), but also have the opportunity to work with collaborators at the University of Florida and the National High Magnetic Field Laboratory in Tallahassee, FL. Student researchers are expected to communicate their research experiences to broad audiences through diverse media. Student researchers also are expected to publish their results and report research findings at professional meetings and to the regional educational community. Funds from this project are used to acquire, operate, and maintain research equipment that is not otherwise available at GSC.