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首页> 外文学位 >Characterization of metalloproteins from Nitrosomonas europaea and Bacillus subtilis.
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Characterization of metalloproteins from Nitrosomonas europaea and Bacillus subtilis.

机译：欧洲亚硝化单胞菌和枯草芽孢杆菌的金属蛋白的表征。

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摘要

Studies were performed on a number of metalloproteins from the bacteria Nitrosomonas europaea and Bacillus subtilis. These proteins were purified and characterized using a combination of spectroscopic and analytical techniques. The first study discussed is focused on the protein YxaG from B. subtilis. In this work, it was demonstrated that this protein is a quercetinase enzyme similar the well characterized copper-containing quercetinase found in species of Aspergillus. Quercetinase is a dioxygenase enzyme that catalyzes the degradation of quercetin, a flavonol with antioxidant properties found in various plants, including onions. Detailed studies revealed that in contrast to the copper-containing system of Aspergillus, the YxaG protein utilizes an iron cofactor, and exhibits a unique inhibitor profile, as well as an electron paramagnetic resonance feature indicating that the iron is present as Fe(II).; In the second study, efforts were initiated to prepare polyhistidine and antibody-epitope fusions of two of the three putative subunits, AmoA and AmoB of the integral membrane protein ammonia monooxygenase from N. europaea. Ammonia monooxygenase catalyzes the first step in nitrification, which is the oxidation of ammonia to hydroxylamine. Mutations were made to plasmids containing sections of the respective genes, and the alterations were introduced back to the genomic copies of the genes through homologous recombination. Cells were then grown, and individual subunits were isolated using immunoprecipitation for the antibody epitopes or metal affinity chromatography for the polyhistidine fusions. Results from the immunoprecipitation studies indicated that the ammonia monooxygenase complex contains multiple copies of the AmoB protein, and that AmoA and AmoB copurify under a variety of conditions.; In the final study, a protein of unknown function present in the periplasm of N. europaea was characterized. This protein, referred to as the small metal binding protein SmbP was found to be induced under increasing levels of copper. Spectroscopic studies have shown that it is capable of binding multiple numbers of a variety of biological metals. The protein is novel, with a seven amino acid motif repeated ten times throughout the primary structure, and may be a new class of metal trafficking proteins. The protein was characterized by a number of spectroscopic techniques, including absorption measurements and electron paramagnetic resonance to characterize two distinct sites with varied properties present in the protein.
机译：对来自 Nitrosomonas europaea 和 Bacillus subtilis 细菌的多种金属蛋白进行了研究。结合光谱学和分析技术对这些蛋白质进行纯化和鉴定。讨论的第一个研究重点是来自 B的蛋白YxaG。枯草。在这项工作中，证明了这种蛋白质是一种槲皮素酶，类似于在曲霉物种中发现的特征明确的含铜槲皮素酶。槲皮素酶是一种双加氧酶，可催化槲皮素的降解，槲皮素是一种在各种植物（包括洋葱）中都具有的抗氧化性能的黄酮醇。详细的研究表明，与曲霉的含铜系统相比，YxaG蛋白利用铁辅因子，并表现出独特的抑制剂谱以及电子顺磁共振特征，表明铁是以Fe（II）的形式存在。在第二项研究中，开始努力从 Neuropaea 制备三个推定亚基中的两个亚基，即膜蛋白氨单加氧酶的AmoA和AmoB的多组氨酸和抗体-表位融合体。氨单加氧酶催化硝化的第一步，即将氨氧化为羟胺。对包含各个基因的片段的质粒进行突变，并且通过同源重组将这些改变引入到基因的基因组拷贝中。然后使细胞生长，并使用用于抗体表位的免疫沉淀或用于多组氨酸融合物的金属亲和色谱法分离各个亚基。免疫沉淀研究的结果表明，氨单加氧酶复合物含有多个拷贝的AmoB蛋白，并且AmoA和AmoB在各种条件下都可以共纯化。在最后的研究中， N周质中存在功能未知的蛋白质。欧罗巴的特征。发现这种蛋白质，称为小金属结合蛋白SmbP，是在铜含量增加的情况下诱导的。光谱研究表明，它能够结合多种生物金属。该蛋白质是新颖的，具有七个氨基酸基序，在整个一级结构中重复十次，并且可能是一类新的金属运输蛋白质。该蛋白质通过多种光谱技术进行了表征，包括吸收测量和电子顺磁共振来表征蛋白质中存在的两个具有不同性质的不同位点。

著录项

  • 作者

    Barney, Brett Michael.;

  • 作者单位

    Arizona State University.;

  • 授予单位 Arizona State University.;
  • 学科 Chemistry Biochemistry.; Biology Microbiology.; Biology Molecular.
  • 学位 Ph.D.
  • 年度 2003
  • 页码 127 p.
  • 总页数 127
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 生物化学;微生物学;分子遗传学;
  • 关键词

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