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The role of cardiolipin in the biogenesis of the Gram-negative bacterial cell envelope

心磷脂在革兰氏阴性细菌细胞包膜生物发生中的作用

基本信息

批准号：
10731444
负责人：
Michael Stephen Trent
金额：
$ 67.71万
依托单位：
UNIVERSITY OF GEORGIA
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-06-01 至 2028-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10731444
关键词：
ATP phosphohydrolase ATP-Binding Cassette Transporters Affinity Antibiotic Resistance Antibiotics Antimicrobial Resistance Automobile Driving Bacteria Bacterial Infections Binding Sites Biochemical Biogenesis Biology Cardiolipins Cell Membrane Permeability Cell Survival Cells Cessation of life Clinical Cytoplasm Data Defect Development Drug Efflux Encapsulated Enzymes Equilibrium Escherichia coli Face Feedback Future Genes Genetic Glycerophospholipids Gram-Negative Bacteria Growth Healthcare Heterogeneity Infection Libraries Lipid A Lipids Lipopolysaccharides Maintenance Membrane Membrane Lipids Modeling Molecular Multi-Drug Resistance Organism Pathway interactions Peptidoglycan Pump Regulation Resistance Role Side Surface System Transmembrane Transport antimicrobial cardiolipin synthase cell envelope combinatorial density design economic cost fighting fitness global health insight lipidome lipophilicity membrane biogenesis mutant new therapeutic target novel novel therapeutics pathogen prevent protein transport stem

项目摘要

Abstract The increasing rise in antibiotic resistance and the diminished discovery of new antimicrobials threatens global healthcare. Of particular concern are Gram-negative pathogens, as these organisms are intrinsically resistant to multiple classes of antibiotics and the discovery of novel drugs targeting these bacteria has remained challenging. The innate resistance of these organisms is provided primarily by their outer membrane (OM), a defining feature of Gram negatives that encapsulates their peptidoglycan layer. Unlike the inner membrane (IM) that is composed solely of glycerophospholipids (GPLs), the OM is asymmetrical with GPLs found in the inner leaflet and lipopolysaccharide (LPS) localized to the outer leaflet. This unique membrane organization affords protection from large polar molecules, as well as lipophilic compounds, creating an impervious barrier. Since the OM is essential, pathways required for its assembly are key targets for antimicrobial design. Currently, there are no antibiotics that directly target OM biogenesis in clinical use and first attempts have proven difficult. Thus, it remains critical to investigate cell envelope biology for future and current antimicrobial design. Recently, we discovered a connection between the GPL cardiolipin (CL) and the synthesis and transport of LPS. E. coli harbors three distinct enzymes that synthesize CL, yet CL is not required for cell viability and is the least abundant of the three major GPLs in Gram negatives. We found LpxM, the enzyme that adds the last acyl chain to the lipid anchor of LPS, to be critical for viability in the absence of clsA. Suppressors of clsA and lpxM synthetic lethality were identified in msbA, a gene that encodes the essential, homodimeric ABC transporter that “flips” LPS across the IM. Multiple pieces of genetic and biochemical data supported a model in which CL enhances MsbA activity driving LPS transport. Also, we observed that single mutants lacking either ClsA, the primary CL synthase, or LpxM have reduced LPS levels. This suggests the cell can “sense” defects in LPS transport at the cytoplasmic face of the IM and slow LPS synthesis to balance OM lipid content. In the current application we will define (i) the functional role of CL in MsbA-dependent LPS transport, (ii) characterize specific MsbA-CL interactions and determine how they impact MsbA activity, (iii) determine if ClsA and MsbA are co-localized in the bacterial cell envelope, and (iv) determine how defects in LPS transport results in feedback inhibition of LPS synthesis. Completion of these Aims will provide novel insights into cell envelope biogenesis and promote the development of novel therapeutics targeting Gram-negative pathogens.

摘要