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Title: Structure and Dynamics of FosA-Mediated Fosfomycin Resistance in Klebsiella pneumoniae and Escherichia coli

Abstract

ABSTRACT Fosfomycin exhibits broad-spectrum antibacterial activity and is being reevaluated for the treatment of extensively drug-resistant pathogens. Its activity in Gram-negative organisms, however, can be compromised by expression of FosA, a metal-dependent transferase that catalyzes the conjugation of glutathione to fosfomycin, rendering the antibiotic inactive. In this study, we solved the crystal structures of two of the most clinically relevant FosA enzymes: plasmid-encoded FosA3 fromEscherichia coliand chromosomally encoded FosA fromKlebsiella pneumoniae(FosA KP). The structure, molecular dynamics, catalytic activity, and fosfomycin resistance of FosA3 and FosA KPwere also compared to those of FosA fromPseudomonas aeruginosa(FosA PA), for which prior crystal structures exist.E. coliTOP10 transformants expressing FosA3 and FosA KPconferred significantly greater fosfomycin resistance (MIC, >1,024 μg/ml) than those expressing FosA PA(MIC, 16 μg/ml), which could be explained in part by the higher catalytic efficiencies of the FosA3 and FosA KPenzymes. Interestingly, these differences in enzyme activity could not be attributed to structural differences at their active sites. Instead, molecular dynamics simulations and hydrogen-deuterium exchange experiments with FosA KPrevealed dynamic interconnectivity between its active sites and a loop structure that extends from the active site of each monomer and traverses the dimer interface. This dimer interface loop is longer and more extendedmore » in FosA KPand FosA3 than in FosA PA, and kinetic analyses of FosA KPand FosA PAloop-swapped chimeric enzymes highlighted its importance in FosA activity. Collectively, these data yield novel insights into fosfomycin resistance that could be leveraged to develop new strategies to inhibit FosA and potentiate fosfomycin activity.« less

Authors:
; ; ; ORCiD logo; ; ; ; ; ; ; ; ; ORCiD logo
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
UNIVERSITYNIH
OSTI Identifier:
1498302
Resource Type:
Journal Article
Journal Name:
Antimicrobial Agents and Chemotherapy
Additional Journal Information:
Journal Volume: 61; Journal Issue: 11; Journal ID: ISSN 0066-4804
Publisher:
American Society for Microbiology
Country of Publication:
United States
Language:
ENGLISH

Citation Formats

Klontz, Erik H., Tomich, Adam D., Günther, Sebastian, Lemkul, Justin A., Deredge, Daniel, Silverstein, Zach, Shaw, JoAnna F., McElheny, Christi, Doi, Yohei, Wintrode, Patrick L., MacKerell, Alexander D., Sluis-Cremer, Nicolas, and Sundberg, Eric J. Structure and Dynamics of FosA-Mediated Fosfomycin Resistance in Klebsiella pneumoniae and Escherichia coli. United States: N. p., 2017. Web. doi:10.1128/AAC.01572-17.
Klontz, Erik H., Tomich, Adam D., Günther, Sebastian, Lemkul, Justin A., Deredge, Daniel, Silverstein, Zach, Shaw, JoAnna F., McElheny, Christi, Doi, Yohei, Wintrode, Patrick L., MacKerell, Alexander D., Sluis-Cremer, Nicolas, & Sundberg, Eric J. Structure and Dynamics of FosA-Mediated Fosfomycin Resistance in Klebsiella pneumoniae and Escherichia coli. United States. doi:10.1128/AAC.01572-17.
Klontz, Erik H., Tomich, Adam D., Günther, Sebastian, Lemkul, Justin A., Deredge, Daniel, Silverstein, Zach, Shaw, JoAnna F., McElheny, Christi, Doi, Yohei, Wintrode, Patrick L., MacKerell, Alexander D., Sluis-Cremer, Nicolas, and Sundberg, Eric J. Tue . "Structure and Dynamics of FosA-Mediated Fosfomycin Resistance in Klebsiella pneumoniae and Escherichia coli". United States. doi:10.1128/AAC.01572-17.
@article{osti_1498302,
title = {Structure and Dynamics of FosA-Mediated Fosfomycin Resistance in Klebsiella pneumoniae and Escherichia coli},
author = {Klontz, Erik H. and Tomich, Adam D. and Günther, Sebastian and Lemkul, Justin A. and Deredge, Daniel and Silverstein, Zach and Shaw, JoAnna F. and McElheny, Christi and Doi, Yohei and Wintrode, Patrick L. and MacKerell, Alexander D. and Sluis-Cremer, Nicolas and Sundberg, Eric J.},
abstractNote = {ABSTRACT Fosfomycin exhibits broad-spectrum antibacterial activity and is being reevaluated for the treatment of extensively drug-resistant pathogens. Its activity in Gram-negative organisms, however, can be compromised by expression of FosA, a metal-dependent transferase that catalyzes the conjugation of glutathione to fosfomycin, rendering the antibiotic inactive. In this study, we solved the crystal structures of two of the most clinically relevant FosA enzymes: plasmid-encoded FosA3 fromEscherichia coliand chromosomally encoded FosA fromKlebsiella pneumoniae(FosAKP). The structure, molecular dynamics, catalytic activity, and fosfomycin resistance of FosA3 and FosAKPwere also compared to those of FosA fromPseudomonas aeruginosa(FosAPA), for which prior crystal structures exist.E. coliTOP10 transformants expressing FosA3 and FosAKPconferred significantly greater fosfomycin resistance (MIC, >1,024 μg/ml) than those expressing FosAPA(MIC, 16 μg/ml), which could be explained in part by the higher catalytic efficiencies of the FosA3 and FosAKPenzymes. Interestingly, these differences in enzyme activity could not be attributed to structural differences at their active sites. Instead, molecular dynamics simulations and hydrogen-deuterium exchange experiments with FosAKPrevealed dynamic interconnectivity between its active sites and a loop structure that extends from the active site of each monomer and traverses the dimer interface. This dimer interface loop is longer and more extended in FosAKPand FosA3 than in FosAPA, and kinetic analyses of FosAKPand FosAPAloop-swapped chimeric enzymes highlighted its importance in FosA activity. Collectively, these data yield novel insights into fosfomycin resistance that could be leveraged to develop new strategies to inhibit FosA and potentiate fosfomycin activity.},
doi = {10.1128/AAC.01572-17},
journal = {Antimicrobial Agents and Chemotherapy},
issn = {0066-4804},
number = 11,
volume = 61,
place = {United States},
year = {2017},
month = {9}
}