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Opposite motion of the Central Helices of efflux pump KmrA is important for its export efficiency

Ying Li1, Honglin Wen2, Xizhen Ge1

  • 1College of Biochemical Engineering, Beijing Union University, Beijing, 100023, China.

Microbial Pathogenesis
|May 13, 2022
PubMed

Insights

The study reveals how amino acid changes in the KmrA efflux pump affect its function in Klebsiella pneumoniae. Disruptions in central helix motion, particularly the S374A substitution, reduce antibiotic export efficiency.

Area of Science:

  • Microbiology
  • Structural Biology
  • Biochemistry

Background:

  • The Major Facilitator Superfamily (MFS) efflux pump KmrA plays a role in antibiotic resistance in Klebsiella pneumoniae.
  • The specific impact of KmrA amino acid variations on its efflux efficiency is not well understood.

Purpose of the Study:

  • To investigate the effect of amino acid substitutions in the KmrA efflux pump on its export efficiency.
  • To elucidate the molecular mechanisms underlying KmrA's function and its regulation.

Main Methods:

  • Bioinformatic analysis of 804 Klebsiella pneumoniae isolates to identify KmrA variants.
  • Microbiological assays (MIC, ethidium bromide efflux) to assess pump activity.
  • Molecular dynamics (MD) simulations to analyze conformational changes and protein dynamics.
  • Analysis of binding pocket volume, salt bridge interactions, and hydrophilic interactions.

Main Results:

  • Four major KmrA variants were identified, with KmrA-A showing inactive efflux.
  • MD simulations revealed that active KmrA variants exhibit opposite motion of central helices, unlike KmrA-A.
  • The S374A substitution in KmrA-A disrupts normal helix motion via enhanced hydrophobic interactions, impacting conformational transitions.
  • Active KmrA's opposite helix motion is linked to a larger binding pocket and stronger water interactions in the inward-facing conformation.

Conclusions:

  • The opposite motion of central helices is crucial for KmrA's export efficiency and conformational transitions.
  • Specific amino acid substitutions, like S374A, can impair KmrA function by altering its dynamic properties.
  • Understanding KmrA's dynamics provides insights into antibiotic resistance mechanisms in Klebsiella pneumoniae.
Keywords:
Antibiotic resistanceEfflux pumpMajor facilitator superfamilyOpposite motion

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