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Allosteric adhesins of enterobacterial pathogens

肠杆菌病原体的变构粘附素

基本信息

批准号：
10626963
负责人：
Rachel E Klevit
金额：
$ 72.02万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-05-24 至 2027-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10626963
关键词：
Adhesions Adhesives Affinity Amino Acids Bacteria Bacterial Adhesins Bacterial Adhesion Binding Binding Proteins Biological Assay Cell Adhesion Cell surface Cells Characteristics Charge Core Protein Cryoelectron Microscopy Crystallography Dissociation Enterobacter Enterobacteriaceae Escherichia coli Escherichia coli Adhesins Eukaryota Family Fimbriae Proteins Fimbrial Adhesins Future Goals Hair Homologous Gene Human Image Kinetics Klebsiella pneumoniae Knowledge Lectin Ligand Binding Ligands Mannose Modeling Molecular Molecular Chaperones Molecular Conformation Multi-Drug Resistance Mutagenesis Mutation Oligosaccharides Pathogenesis Pathway interactions Physiological Pilum Positioning Attribute Property Protein Dynamics Proteins Proteus mirabilis Receptor Cell Rod Salmonella Shapes Side Solvents Structure Surface Testing Tissues Uropathogenic E. coli Vaccines Virulence Factors Work X-Ray Crystallography analog appendage conformational conversion conformer design fimbria human pathogen hydrophilicity insight mechanical force molecular dynamics mutant pathogen pathogenic bacteria pragmatic implementation protein structure shear stress small molecule inhibitor success sugar tool

项目摘要

Abstract This proposal seeks to identify potential allosteric properties in adhesins of human enterobacterial pathogens - Escherichia coli, Klebsiella pneumoniae/oxytoca, Enterobacter spp, Proteus mirabilis, and Salmonella – that are assembled via a chaperone-usher pathway (CUP). To date, only the mannose-specific, type 1 fimbrial adhesin of E. coli, FimH, has been demonstrated to be an allosteric protein that can exist in alternative functional (active/inactive) conformations. This property allows bacteria that contain FimH as part of hair-like surface appendages, fimbriae or pili, to bind ligand presented on host cells rapidly from an inactive conformation and to remain bound for very long lifetimes under shear force by transiting to an active conformation. The long-lived (slow dissociation) binding involves formation of so-called `catch-bonds' that can be activated and become stronger under tensile mechanical force and involve an allosteric switch. To date no other bacterial adhesin has been demonstrated to be allosteric and to exist in alternative functional (active/inactive) conformations. To identify other adhesins that work via similar mechanisms, we will focus on adhesins that are part of fimbriae or pili and belong to the same CUP structural class as FimH. We recently identified a set of aliphatic or aromatic residues that act as molecular toggles that control the allosteric switch between active and inactive conformations by switching their orientation between the protein core and surface. It is possible to stabilize either active or inactive conformation of the adhesin by “surface locking” such toggles through substitution to hydrophilic charged residues. We will use putative analogs of the FimH toggles to identify the existence of allosteric states in other CUP adhesins that are homologous or non-homologous to FimH, using mutagenesis, various functional assays, and three types of structural analysis – NMR, X-ray crystallography, and cryo-EM. Success of our studies will contribute to understanding of general mechanisms of bacterial adhesion to host cells and, ultimately, to the design of optimized vaccines and small molecule inhibitors. If certain adhesins are found to be allosteric, in-depth analysis of their physiologically-relevant structure/functional properties and significance for pathogenesis as well as practical implementation of the findings will be the focus of future studies.

摘要