The role of BamA in the biogenesis of beta-barrel membrane proteins

BamA 在 β-桶膜蛋白生物合成中的作用

• 批准号: 8767875
• 负责人: Nicholas Noinaj
• 金额: $ 15.92万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-15 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8767875
• 关键词: Academia; Agonist; Bacteria; Binding; Biochemical; Biogenesis; Cause of Death; Cell Survival; Cessation of life; Communicable Diseases; Complex; Country; Cytoplasm; Cytoprotection; Detergents; Development; Electron Microscopy; Faculty; Funding; Future; G Protein-Coupled Receptor Genes; Goals; Gram-Negative Bacteria; Hemophilus ducreyi; Human; Immune response; Infectious Agent; Institution; Knowledge; Label; Learning; Length; Lipoproteins; Manuscripts; Mediating; Membrane; Membrane Proteins; Methods; Molecular; Molecular Chaperones; Multi-Drug Resistance; Names; Nature; Neisseria; Neisseria gonorrhoeae; Neurotensin; Neurotensin Receptors; Neurotransmitters; Nutrient; Peptides; Peptidoglycan; Phase; Proteins; Publishing; Reporting; Research; Role; Route; Signal Transduction; Site; Spectrum Analysis; Structure; Surface; Technology; Transferrin; Virulence; Work; X-Ray Crystallography; base; beta barrel; career; crosslink; fascinate; insight; interest; member; membrane biogenesis; mimetics; pandemic disease; pathogenic bacteria; periplasm; protein folding; public health relevance; receptor; resistant strain; success

DESCRIPTION (provided by applicant): Infectious diseases cause widespread sickness throughout the world each year and are the second leading cause of death, particularly in underdeveloped countries. And with the emergence of multi-drug resistance strains, the necessity for new, more effective, and more sustainable therapies is immediate and vital to protect against any future pandemics. My studies will provide crucial insight into the biogenesis of surface receptors and proteins that assist pathogenic bacteria in their virulence. This knowledge will significantly assist in the development of better therapies against these infectious agents. Gram-negative bacteria contain an inner and outer membrane. The outer membrane contains a host of beta-barrel proteins commonly called outer membrane proteins (OMPs), which serve essential functions in cargo transport, signaling, and bacterial virulence. In Gram-negative bacteria, it is known that OMPs are synthesized in the cytoplasm and then transported across the inner membrane into the periplasm via a Sec translocon. Once in the periplasm, chaperones guide the nascent OMPs across the periplasm and peptidoglycan to the inner surface of the outer membrane. Here, the nascent OMPs are recognized by a complex known as the BAM complex which folds and inserts the new OMPs into the outer membrane. Exactly how the BAM complex is able to accomplish its function remains unknown. However, we do know that the BAM complex consists of five subunits named BamA (an OMP itself), BamB, BamC, BamD, and BamE, which are all lipoproteins. Studies have shown that BamA and BamD are absolutely essential for cell viability and OMP biogenesis. Our lab and others have reported the structures of BamB, BamC, BamD, BamE and a large portion of the periplasmic domain of BamA, providing initial insight into how the BAM complex may function. However, even with these structures being known, the mechanism for how the BAM complex recognizes, folds, and inserts nascent OMPs into the outer membrane remains elusive, largely due to the lack of a full length BamA structure and complexes with BamA. Recently, I determined the crystal structures of a truncated BamA construct from Haemophilus ducreyi to 2.9 ¿ and of a full length BamA construct from Neisseria gonorrhea to 3.2 ¿. In my proposed studies, I aim to build on this recent success to use X-ray crystallography to determine structures of BamA in complex with BamB-E, to use DEER spectroscopy and crosslinking to characterize the conformational dynamics of BamA, and to use crosslinking to explore the interactions between nascent OMPs and BamA and the other BAM components, with my goal being understand the functional role of BamA within the BAM complex. I have a strong background in X-ray crystallography and during my postdoctoral studies, have added a strong background in working with and crystallizing membrane proteins using the latest technologies such as new stabilizing detergents, bicelles, and lipidic cubic phase (LCP) methods. Recently, I solved the structures of two important surface proteins from pathogenic Neisseria and this work was published in Nature as a full research article (March 2012). For this manuscript, I also solved the structure of diferric human transferrin which research groups have been trying to solve for decades without success. More recently, I also solved the crystal structure of the agonist bound neurotensin receptor NTSR1, a GPCR responsible for binding neurotensin and other neurotransmitters. This work was published in Nature as well in October 2012 (Research Article). My current efforts are focused on studying BamA of the BAM complex and I will take this project with me as a tenure-track faculty member in academia. And with this, my most recent results, the crystal structures of BamA (H. ducreyi and N. gonorrhea), were also recently published in Nature as a full research article as well (Sept 2013). My long-term goal is to have my own research lab as a faculty member at a Research I academic institution where I can continue my research interests. This has been my lifelong ambition and while funding for academic research is more competitive now than ever, I remain dedicated to a career in research with aspirations that I will be able to establish a research group that significantly advances our current understanding of beta-barrel membrane proteins in both bacteria and humans.

描述（由申请人提供）：传染病每年在世界各地引起广泛的疾病，是第二大死亡原因，特别是在不发达国家。随着耐多药菌株的出现，迫切需要新的、更有效和更可持续的治疗方法，以防止未来的任何大流行。我的研究将为表面受体和蛋白质的生物发生提供重要的见解，这些受体和蛋白质有助于致病菌的毒力。这一知识将大大有助于开发更好的治疗这些传染性疾病的方法