Cytochrome C Biogenesis

细胞色素C生物发生

• 批准号: 7267433
• 负责人: Robert G. Kranz
• 金额: $ 29.15万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-09-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7267433
• 关键词: ATP Hydrolysis; ATP-Binding Cassette Transporters; Address; Aerobic; Affinity; Arabidopsis; Archaeoglobus; Avidity; Bacillus (bacterium); Binding; Binding Sites; Biochemical Genetics; Bioenergetics; Biogenesis; C-terminal; Caenorhabditis; Cell membrane; Cells; Chimeric Proteins; Chlamydomonas; Chloroplasts; Complex; Cryptosporidium; Cysteine; Cytochrome c Group; Cytochromes; Cytoplasm; Defect; Deinococcus; Dependence; Engineering; Enzymes; Escherichia; Escherichia coli; Euglena; Genes; Genus Mycobacterium; Gram-Positive Bacteria; Growth; Hela Cells; Helicobacter; Heme; Hemeproteins; Hereditary Disease; Human; Invertebrates; Life; Ligase; Ligation; Literature; Membrane; Membrane Proteins; Metals; Mitochondria; Molecular; Molecular Chaperones; Mus; Myxococcus; N-terminal; Nature; Nomenclature; Object Attachment; Organism; Oxidation-Reduction; Paramecium; Pathway interactions; Plants; Plasmodium; Porphyrins; Process; Production; Prokaryotic Cells; Property; Proteins; Proteobacteria; Protoporphyrins; Protozoa; Recombinants; Research Personnel; Rhodobacter; Role; Saccharomyces; Side; Signal Transduction; Surface; Synechocystis; System; Testing; Tetrahymena; Time; Triticum; Volvox; analog; antimicrobial drug; base; cofactor; cytochrome C synthetase; cytochrome c; ferrochelatase; frontier; heme a; in vivo; inhibitor/antagonist; pathogen; periplasm; porin; protein function; reconstitution; success; thioether

DESCRIPTION (provided by applicant): Cytochromes are heme proteins essential for the aerobic and anaerobic growth of most organisms, including human pathogens. Relatively recently it has become clear that dedicated assembly pathways and factors are crucial for cytochrome biogenesis. The assembly of c-type cytochromes occurs by one of three pathways, called systems I, II, and III. Systems I and II have nine and four assembly factors (membrane proteins) respectively, while system III uses a single enzyme called cytochrome c heme lyase. Defects in system III result in certain human genetic diseases. Because only prokaryotes, plants, and protozoa use systems I or II, these pathways represent potential targets for antimicrobial agents. The c-type cytochromes possess heme that is covalently ligated to the apocytochrome at two cysteines, which must be reduced for attachment to the heme. Since assembly of c-type cytochromes occurs at the outer surface of the inner membrane, cells must deliver reduced heme, synthesized inside the cell, to the secreted unfolded apocytochrome. In this application we take advantage of our previous success for reconstituting all three systems in recombinant Escherichia coli to study the in vivo concentrations and features of heme that are optimal for each system. The heme delivery pathways will be established. Furthermore, the protein(s) required for the final cytochrome c synthetase (ligase) activity of each system will be purified and characterized for heme and apocytochrome c ligation properties, binding, and mechanisms. Based on our findings that certain metal porphyrins are specific inhibitors of the systems I and II pathways, other porphyrin analogs will be tested for their ability to be ligated to apocytochrome c (or to inhibit biogenesis), further delineating the properties of heme that are critical for ligation. In summary, this study will address which protein(s) constitute the cytochrome c synthetases, how heme is delivered, what concentrations and features of heme are required for each system. Results will help unravel why three very different pathways have emerged in nature and mechanisms to target them.

描述（由申请人提供）：细胞色素是大多数生物（包括人病原体）的有氧和厌氧生长所必需的血红素蛋白。相对较新的很明显，专用的组装途径和因素对于细胞色素生物发生至关重要。 C型细胞色素的组装由三种途径之一，称为系统I，II和III。系统I和II分别具有九个和四个组装因子（膜蛋白），而系统III使用一种称为细胞色素C血红素裂解酶的单个酶。系统III的缺陷导致某些人类遗传疾病。因为只有原核生物，植物和原生动物使用系统I或II，所以这些途径代表了抗菌剂的潜在靶标。 C型细胞色素具有血红素，该血红素在两个半胱氨酸处共价连接到启示剂，必须降低以固定在血红素上。由于C型细胞色素的组装发生在内膜的外表面，因此细胞必须递送血红素，在细胞内部合成的血红素，向分泌的展开的脱落式染色组。在此应用中，我们利用了以前的成功来重组大肠杆菌中的所有三个系统，以研究针对每个系统最佳的血红素的体内浓度和特征。将建立血红素递送途径。此外，每个系统的最终细胞色素c合成酶（连接酶）活性所需的蛋白质将被纯化，并针对血红素和启示膜C结扎特性，结合和机制进行表征。根据我们的发现，某些金属卟啉是系统I和II途径的特定抑制剂，将测试其他卟啉类似物的能力，以使其与脱染色剂C（或抑制生物发生）结扎的能力，进一步描述了血红素的特性，这些性质是至关重要的。进行连接。总而言之，这项研究将介绍哪种蛋白质构成细胞色素C合成酶，血红素的输送方式，每个系统所需的血红素的浓度和特征。结果将有助于解散为什么在性质和机制中出现了三种截然不同的途径来瞄准它们。