QUINOENZYMES--BIOGENESIS STRUCTURE AND FUNCTION

在过去的五年里，一种新的蛋白质被描述出来， 命名为醌蛋白。 这些蛋白质含有它们的辅因子 在它们的多肽骨架中，揭示了一种新的策略， 生物学中氧化还原辅因子的产生。 结合使用 化学、生物化学和分子生物学方法， 我们会继续研究这些系统。 详细调查了 铜胺氧化酶中辅因子topa quinone（TPQ）的生物发生 (CAO的）计划，目的是了解如何CAO的 能够催化“自我加工”。 迄今已有的数据 暗示CAO的活性位点铜在辅因子生物成因和 催化周转 虽然正式成为曹操家族的一员， 赖氨酰氧化酶（LO）由于其减小的大小而不同， 缺乏显著序列同源性。 初步数据显示， 实验室表明，虽然LO含有醌结构， 与以前在其他蛋白质中看到的结构不同。 大 一些实验计划，涉及表征活性 位点衍生肽及其模型的合成与比较研究 化合物. 四种克隆和测序的真核生物的序列同源性 CAO’s表明了一个保守的结构基序朝向C-末端， 蛋白质;该结构基序包含TPQ共有序列 和铜的组氨酸配体。 酵母胺的基因 来自多形汉逊酵母（YAO）的氧化酶将被修饰用于表达 C-末端结构域，目的是确定是否辅因子 生物发生可以发生在这个区域内。 目前正在努力， 将继续获得YAO的X射线结构，并最终 延伸到YAO的突变形式。 解决方案研究，侧重于 胺氧化酶从牛血浆和姚，将解决一些 这些问题包括活性位点共有序列的作用， 确定底物特异性、活性位点碱基的性质 催化底物氧化，和半的化学机制， 将分子氧还原成过氧化氢的反应。 给定 其广泛的生理功能，包括氧化 从血液中去除生物胺并交联 胶原蛋白和弹性蛋白，正在研究的quinoproteins直接相关 对人类健康的影响。

During the past five years a new class of proteins has been described, designated quinoproteins. These proteins, which contain their cofactor within their polypeptide backbone, reveal a novel strategy for the generation of redox cofactors in biology. Utilizing a combination of chemical, biochemical and molecular biological approaches, multifaceted studies of these systems will be pursued. A detailed investigation of the biogenesis of the cofactor topa quinone (TPQ) in the copper amine oxidases (CAO's) is planned, with the goal of understanding how the CAO's are capable of catalyzing "self processing". Data available thus far implicate the active site copper of CAO's in both cofactor biogenesis and catalytic turnover. Although formally a member of the CAO family of proteins, lysyl oxidase (LO) differs by virtue of its reduced size and lack of significant sequence homology. Preliminary data from this laboratory indicate that while LO contains a quino-structure, this is different from structures seen previously in other proteins. A large number of experiments are planned, involving characterization of active site derived peptides and the synthesis and comparative study of model compounds. Sequence homology among four cloned and sequenced eukaryotic CAO's indicates a conserved structural motif toward the C-terminus of protein; this structural motif contains both the TPQ consensus sequence and the putative histidine ligands to copper. The gene for yeast amine oxidase from Hansenula polymorpha (YAO) will be modified for expression of the C-terminal domain, with the goal of determining whether cofactor biogenesis can occur within this domain. Efforts currently underway, to obtain an X-ray structure for YAO will be continued and eventually extended to mutant forms of YAO. Solution studies, focused on both an amine oxidase from bovine plasma and YAO, will address a number of questions which include the role of the active site consensus sequence in determining substrate specificity, the nature of the active site base catalyzing substrate oxidation, and the chemical mechanism of the half reaction involving reduction of dioxygen to hydrogen peroxide. Given their wide ranging physiologic functions which include the oxidative removal of biogenic amines from the blood stream and the cross-linking of collagen and elastin, the quinoproteins under study are directly related to human health.