Unexpected parallels between SaPI replication initiators and conserved SCC ORFs

SaPI 复制启动子和保守的 SCC ORF 之间的意外相似之处

• 批准号: 9001253
• 负责人: PHOEBE A RICE
• 金额: $ 19.75万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-02-01 至 2018-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9001253
• 关键词: ATP phosphohydrolase; ATPase Domain; Address; Adenylyl Imidodiphosphate; Antibiotic Resistance; Appearance; Binding; Biochemical; Biochemistry; Biological Models; Biology; Chromosomes; Complex; Crystallization; Crystallography; DNA; DNA biosynthesis; Data; Data Set; Drug Design; Elements; Enzymes; Event; Excision; Family; Future; Gene Structure; Genes; Genome; Genomic Islands; Homologous Gene; Horizontal Gene Transfer; Housekeeping Gene; Literature; Medical; Methicillin Resistance; Mobile Genetic Elements; Molecular Biology; Morphology; Open Reading Frames; Pathogenesis; Pathogenicity Island; Pattern; Phase; Proteins; Public Health; Replication Initiation; Replication Origin; Resistance; Site; Staphylococcus aureus; Structure; Superantigens; Testing; Thinking; Toxin; Variant; Work; base; biochemical tools; clinical practice; ds-DNA; helicase; in vitro activity; in vivo; melting; methicillin resistant Staphylococcus aureus; prevent; public health relevance; recombinase; resistance gene

 DESCRIPTION (provided by applicant): This project will investigate a new hypothesis regarding how the mobile genetic element (SCCmec) that carries methicillin resistance spreads among S. aureus strains: We propose that it encodes previously unsuspected functionality for its own replication, which would enhance the efficiency of any horizontal gene transfer mechanism. MRSA (methicillin resistant S. aureus) is a serious and growing public health problem. The recombinases that integrate and excise the SCCmec element into and out of the host genome have been identified and are under study in our lab. However, nothing is known about the events between excision from one genome and appearance in another. While analyzing the conserved ORFs that flank recombinase genes, we realized that some of are likely to have functions in DNA replication. This proposal focuses on a large putative ATPase (Cch) that we discovered is related to the known replication initiation proteins encoded by S. aureus pathogenicity islands (SaPIs). Our preliminary structural and biochemical data suggest that the SCCmec protein in question (Cch) is a self-loading helicase whose closest homologs in the ATPase domain are, surprisingly, the eukaryotic and archaeal replicative MCM helicases. Thus we have determined the first structure of that type of helicase in the active ring form. This work will change how people think about SCCmec elements and how they spread methicillin resistance and will provide new model systems for understanding mechanisms of self-loading and MCM-like helicases. Aim 1: Structural understanding of Cch Aim 2: How structurally similar / different are the subtypes of SaPI / SCC replication initiator? Aim 3: What are the biochemical activities of Cch and SaPI3 rep?

 描述（由申请人提供）：本项目将研究一个新的假设，即携带甲氧西林耐药性的移动的遗传元件（SCCmec）如何在沙门氏菌中传播。金黄色葡萄球菌菌株：我们认为它编码了以前未被怀疑的自身复制功能，这将提高任何水平基因转移机制的效率。MRSA（耐甲氧西林S.金黄色葡萄球菌）是一个严重的和日益严重的公共卫生问题。将SCCmec元件整合到宿主基因组中并从宿主基因组中切除的重组酶已经被鉴定，并且正在我们的实验室中进行研究。然而，对于从一个基因组切除到出现在另一个基因组之间的事件，我们一无所知。在分析重组酶基因侧翼的保守ORF时，我们意识到其中一些可能在DNA复制中起作用。本研究的重点是我们发现的一个大的ATP酶（Cch），它与已知的由S.金黄色葡萄球菌致病岛（SaPI）。我们的初步结构和生化数据表明，SCCmec蛋白的问题（Cch）是一个自加载解旋酶的ATP酶结构域中最接近的同系物，令人惊讶的是，真核生物和古细菌的复制MCM解旋酶。因此，我们已经确定了活性环状解旋酶的第一个结构。这项工作将改变人们对SCCmec元件的看法，以及它们如何传播甲氧西林耐药性，并将为理解自加载和MCM样解旋酶的机制提供新的模型系统。目的1：Cch的结构理解目的2：SaPI / SCC复制启动子的亚型在结构上有多相似/不同？目的3：Cch和SaPI 3 rep的生化活性是什么？