Deciphering Multidrug Efflux: Substrate Interactions and Transport Mechanisms in PatAB and NorM-VC
type: CreativeWork
description: Multidrug efflux transporters are ubiquitously expressed in all kingdoms of life. In bacteria, they participate in several innate physiological functions to aid survival and adaptation. Given their crucial roles in conferring antimicrobial resistance (AMR), the understanding of the structural and functional aspects of these transporters is essential for developing strategies to combat AMR. Despite recent advances in structural biology, several key aspects of these transporters remain unclear. These include the molecular details of how they recognise, interact with, and transport substrates; the detailed mechanisms of energy coupling to substrate transport; and the functional implications of the structural asymmetry in heterodimeric ATP-binding cassette (ABC) multidrug transporters. To address these gaps, this work focuses on two distinct types of multidrug efflux transporters: the ABC transporter PatAB from Streptococcus pneumoniae and the multidrug and toxic compound extrusion (MATE) transporter NorM-VC from Vibrio cholerae. First, the study on PatAB revealed two classes of transported substrates based on their effects on nucleotide hydrolysis activity. While most substrates had little to no impact on nucleotide hydrolysis activity, a group of inhibitory substrates were identified, including ethidium, propidium, novobiocin, and coumermycin A1, which significantly decrease nucleotide hydrolysis activity at low concentrations through non-competitive inhibition. Analyses of nucleotide binding confirmed that these inhibitors do not compete with nucleotide binding but that they bind to allosteric sites on PatAB, possibly at the interface between the nucleotide-binding domains (NBDs) or near the coupling helices at the interface between NBDs and transmembrane domains (TMDs). Next, mutations in the consensus and degenerate D-loops of the NBDs were introduced to investigate the complex coupling between nucleotide hydrolysis and substrate transport of PatAB. The consensus D…
abstract: Multidrug efflux transporters are ubiquitously expressed in all kingdoms of life. In bacteria, they participate in several innate physiological functions to aid survival and adaptation. Given their crucial roles in conferring antimicrobial resistance (AMR), the understanding of the structural and functional aspects of these transporters is essential for developing strategies to combat AMR. Despite recent advances in structural biology, several key aspects of these transporters remain unclear. These include the molecular details of how they recognise, interact with, and transport substrates; the detailed mechanisms of energy coupling to substrate transport; and the functional implications of the structural asymmetry in heterodimeric ATP-binding cassette (ABC) multidrug transporters. To address these gaps, this work focuses on two distinct types of multidrug efflux transporters: the ABC transporter PatAB from Streptococcus pneumoniae and the multidrug and toxic compound extrusion (MATE) transporter NorM-VC from Vibrio cholerae. First, the study on PatAB revealed two classes of transported substrates based on their effects on nucleotide hydrolysis activity. While most substrates had little to no impact on nucleotide hydrolysis activity, a group of inhibitory substrates were identified, including ethidium, propidium, novobiocin, and coumermycin A1, which significantly decrease nucleotide hydrolysis activity at low concentrations through non-competitive inhibition. Analyses of nucleotide binding confirmed that these inhibitors do not compete with nucleotide binding but that they bind to allosteric sites on PatAB, possibly at the interface between the nucleotide-binding domains (NBDs) or near the coupling helices at the interface between NBDs and transmembrane domains (TMDs). Next, mutations in the consensus and degenerate D-loops of the NBDs were introduced to investigate the complex coupling between nucleotide hydrolysis and substrate transport of PatAB. The consensus D…
author: Hsieh, Pei-Yu
language: eng
keywords: ABC transporters, Antimicrobial resistance, MATE transporters, Membrane transporters, Multidrug efflux transporters
license: https://www.repository.cam.ac.uk/bitstreams/399aee00-cd32-43b0-8588-facd2a716474/download
license: http://purl.org/NET/rdflicense/allrightsreserved
date published: 2024-09-30
date modified: 2026-07-24T01:33:25Z
provider: Cambridge University
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Cambridge

PropertyValue
property ID: DOI
value: 10.17863/CAM.115307