AcrB drug-binding pocket substitution confers clinically relevant resistance and altered substrate specificity.
The incidence of multidrug-resistant bacterial infections is increasing globally and the need to understand the underlying mechanisms is paramount to discover new therapeutics. The efflux pumps of Gram-negative bacteria have a broad substrate range and transport antibiotics out of the bacterium, conferring intrinsic multidrug resistance (MDR). The genomes of pre- and posttherapy MDR clinical isolates of Salmonella Typhimurium from a patient that failed antibacterial therapy and died were sequenced. In the posttherapy isolate we identified a novel G288D substitution in AcrB, the resistance-nodulation division transporter in the AcrAB-TolC tripartite MDR efflux pump system. Computational structural analysis suggested that G288D in AcrB heavily affects the structure, dynamics, and hydration properties of the distal binding pocket altering specificity for antibacterial drugs. Consistent with this hypothesis, recreation of the mutation in standard Escherichia coli and Salmonella strains showed that G288D AcrB altered substrate specificity, conferring decreased susceptibility to the fluoroquinolone antibiotic ciprofloxacin by increased efflux. At the same time, the substitution increased susceptibility to other drugs by decreased efflux. Information about drug transport is vital for the discovery of new antibacterials; the finding that one amino acid change can cause resistance to some drugs, while conferring increased susceptibility to others, could provide a basis for new drug development and treatment strategies.
Antimicrobials Research Group, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Microbiology and Infection, The University of Birmingham, Birmingham B15 2TT, United Kingdom;
Jessica M A
JM
Blair
Antimicrobials Research Group, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Microbiology and Infection, The University of Birmingham, Birmingham B15 2TT, United Kingdom;
Vassiliy N
VN
Bavro
Antimicrobials Research Group, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Microbiology and Infection, The University of Birmingham, Birmingham B15 2TT, United Kingdom;
Vito
V
Ricci
School of Engineering and Science, Jacobs University Bremen, 28759 Bremen, Germany; and.
Niraj
N
Modi
Department of Physics, University of Cagliari, 09042 Monserrato, Italy.
Pierpaolo
P
Cacciotto
School of Engineering and Science, Jacobs University Bremen, 28759 Bremen, Germany; and.
Ulrich
U
Kleinekathӧfer
Department of Physics, University of Cagliari, 09042 Monserrato, Italy.
Paolo
P
Ruggerone
Department of Physics, University of Cagliari, 09042 Monserrato, Italy.
Attilio V
AV
Vargiu
Antimicrobials Research Group, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Microbiology and Infection, The University of Birmingham, Birmingham B15 2TT, United Kingdom;
Alison J
AJ
Baylay
Antimicrobials Research Group, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Microbiology and Infection, The University of Birmingham, Birmingham B15 2TT, United Kingdom;
Helen E
HE
Smith
Antimicrobials Research Group, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Microbiology and Infection, The University of Birmingham, Birmingham B15 2TT, United Kingdom;
Yvonne
Y
Brandon
Antimicrobials Research Group, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Microbiology and Infection, The University of Birmingham, Birmingham B15 2TT, United Kingdom;
David
D
Galloway
Antimicrobials Research Group, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Microbiology and Infection, The University of Birmingham, Birmingham B15 2TT, United Kingdom; l.j.v.piddock@bham.ac.uk.
Laura J V
LJ
Piddock
D019943:Amino Acid Substitution; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D002939:Ciprofloxacin / Q000494:pharmacology; D004317:Doxorubicin / Q000737:chemistry / Q000378:metabolism; D024901:Drug Resistance, Multiple, Bacterial / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics / Q000302:isolation & purification; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D056084:Genetic Fitness; D016680:Genome, Bacterial; D006801:Humans; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D008911:Minocycline / Q000494:pharmacology; D008958:Models, Molecular; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics* / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D019779:Salmonella enterica / Q000187:drug effects / Q000235:genetics / Q000302:isolation & purification; D013379:Substrate Specificity / Q000187:drug effects; D014867:Water / Q000737:chemistry
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D002939:Ciprofloxacin; D004317:Doxorubicin; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D008911:Minocycline; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D014867:Water; C083496:AcrB protein, E coli; C474926:AcrB protein, Salmonella enterica
AcrB; antimicrobial resistance; efflux; whole genome sequencing