STRUCTURAL BASIS OF AMINORABINOSE BIOSYNTHESIS LINKED TO POLYMYXIN RESISTANCE

与多粘菌素抗性相关的氨基阿拉伯糖生物合成的结构基础

• 批准号: 10017248
• 负责人: Vasileios I Petrou
• 金额: $ 24.9万
• 依托单位: RBHS-NEW JERSEY MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-13 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10017248
• 关键词: Active Sites; Address; Anabolism; Antibiotic Resistance; Antibiotics; Antimicrobial Resistance; Award; Bacteria; Bacterial Genome; Binding; Biology; Biomedical Technology; Catalysis; Cations; Chemicals; Chimera organism; Collection; Communities; Cryoelectron Microscopy; Crystallization; Development Plans; Docking; Dolichol Phosphates; Educational process of instructing; Electron Microscopy; Electron Spin Resonance Spectroscopy; Electrostatics; Environment; Enzymes; Eukaryota; Faculty; Family; Future; Genes; Genomic approach; Goals; Gram-Negative Bacteria; Grant; Health; Host Defense; Human; Incidence; Infection; Knowledge; Laboratories; Lead; Ligands; Link; Lipid A; Lipid Binding; Lipids; Lipopolysaccharides; Location; Membrane; Membrane Lipids; Membrane Proteins; Mentors; Methodology; Modification; Molecular; Molecular Conformation; Monosaccharides; Multi-Drug Resistance; Mutagenesis; Names; Orthologous Gene; Pathway interactions; Peptides; Phase; Polymyxin Resistance; Polymyxins; Polysaccharides; Positioning Attribute; Principal Investigator; Probability; Professional Competence; Reaction; Research; Research Proposals; Resistance development; Resort; Resources; Salmonella; Site; Structure; Therapeutic; Training; Transferase; United States National Institutes of Health; Universities; World Health Organization; Writing; X-Ray Crystallography; analog; antimicrobial drug; career; career development; design; enzyme structure; experience; flexibility; global health; glycosyltransferase; inorganic phosphate; member; multidisciplinary; particle; periplasm; programs; responsible research conduct; screening; structural biology; structural genomics; sugar; technology development; tenure track

Project Summary/Abstract The research proposal outlined in this application is consisted of two components designed to facilitate the transition of the principal investigator (PI) to an independent research career, a career development plan and a research plan. The PI has a multidisciplinary background, and through this proposal, seeks to finalize his training in Structural Biology of Membrane Proteins. The career development plan comprises a structured educational experience for the first two years of the award (K99 phase) that includes training in Electron Microscopy and Electron Paramagnetic Resonance (EPR) Spectroscopy for the study of Membrane Proteins, and enhancement of career skills, such as grant writing, laboratory management, teaching and responsible conduct of research. It includes a clear and actionable plan for identifying and successfully competing for an independent tenure-track faculty position by the end of the K99 phase. The PI has assembled a multi- disciplinary team of mentors, advisors and collaborators that will oversee and guide his training, research program and transition to independence. The research plan spans both the mentored (K99) and independent (R00) phases of the award. It involves mechanistic studies of the aminoarabinose sugar biosynthetic pathway in Gram-negative bacteria that is linked to resistance to polymyxin-class antibiotics, our last line of defense against multi-drug resistant infections. The research program for the K99 phase aims to build on knowledge gained from a recent solution structure of an enzyme in the aminoarabinose pathway, named ArnT, by the PI and his colleagues. The independent (R00) research program then aims to extend the structural studies to other transmembrane enzymes of the aminoarabinose pathway, working towards a complete structural description and functional characterization of the pathway. The core questions that the research program aims to address are: (1) How does the enzyme ArnT accommodate its two lipidic substrates within the context of its fold? (2) What are the molecular determinants that impart stereospecificity to some members of the ArnT enzyme family but not others? (3) How is the ArnT enzyme changing during substrate binding and catalysis? (4) How do the other transmembrane enzymes in the pathway look and how do they accomplish their functions? (5) How do other transmembrane enzymes in the pathway accommodate the lipidic sugar carrier undecaprenyl phosphate? To address these questions, the PI has put together a comprehensive research plan that utilizes state-of-the-art methodologies. Moreover, the plan includes some technology development aimed at extending the capabilities of electron microscopy within the context of the research program, and which, upon completion of the program, may be applicable to other research programs. The proposed studies for the K99 phase will largely take place in the prominent environment of Columbia University, which harbors a vibrant structural biology community. This environment is certain to facilitate the PI in the successful completion of the proposed research program and achieving his goal of transitioning into an independent research career.

项目总结/摘要 本申请中概述的研究建议由两个组成部分组成，旨在促进 主要研究者（PI）向独立研究职业的过渡，职业发展计划和 研究计划。PI具有多学科背景，并通过这一建议，寻求完成他的 膜蛋白结构生物学培训。职业发展计划包括结构化的 该奖项前两年（K99阶段）的教育经验，包括电子培训 用于膜蛋白研究的显微镜和电子顺磁共振（EPR）光谱， 和职业技能的提高，如拨款写作，实验室管理，教学和责任 进行研究。它包括一个明确的和可操作的计划，以确定和成功地竞争， 在K99阶段结束时获得独立的终身教职。私家侦探组织了一个多- 由导师、顾问和合作者组成的纪律团队，将监督和指导他的培训、研究 计划和过渡到独立。该研究计划涵盖了指导（K99）和独立 (R00)奖的阶段。它涉及氨基阿拉伯糖糖生物合成途径的机理研究 在革兰氏阴性菌中，它与对多粘菌素类抗生素的耐药性有关，这是我们最后的防线， 对抗多重抗药性感染。K99阶段的研究计划旨在建立在知识的基础上 PI从最近获得的氨基阿拉伯糖途径中一种酶的溶液结构中获得，该酶名为ArnT 和他的同事独立（R 00）研究计划的目的是将结构研究扩展到 氨基阿拉伯糖途径的其他跨膜酶，致力于完整的结构 途径的描述和功能表征。该研究计划的核心问题 要解决的问题是：（1）酶ArnT如何在其作用的背景下适应其两种底物， 弃牌？(2)什么是分子决定因素，赋予立体特异性的一些成员的ArnT 酶家族而不是其他？(3)ArnT酶在底物结合和催化过程中是如何变化的？ (4)途径中的其他跨膜酶是什么样子的，它们是如何完成它们的功能的？ 功能？(5)通路中的其他跨膜酶如何适应糖载体 磷酸十一异戊二烯酯？为了解决这些问题，PI制定了一项全面的研究计划 使用最先进的方法此外，该计划还包括一些技术开发， 在研究计划的范围内扩展电子显微镜的能力，并且， 完成该计划后，可适用于其他研究计划。拟议的研究 K99阶段将主要在哥伦比亚大学的优越环境中进行，这里充满了活力 结构生物学社区这样的环境一定会促进PI在顺利完成 他提出了研究计划，并实现了向独立研究生涯过渡的目标。