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Phage display-derived inhibitor of the essential cell wall biosynthesis enzyme MurF

Catherine Paradis-Bleau1, Adrian Lloyd, François Sanschagrin

  • 1Département de Biologie Médicale, Université Laval, Sainte-Foy, Québec G1K 7P4, Canada. Catherine_Paradis-Bleau@hms.harvard.edu

BMC Biochemistry
|December 23, 2008
PubMed
Abstract

Insights

We identified MurFp1, a novel peptide inhibitor targeting the MurF enzyme in Pseudomonas aeruginosa. This peptide interferes with D-Ala-D-Ala utilization, offering a new strategy against antibiotic-resistant bacteria.

Area of Science:

  • Microbiology
  • Biochemistry
  • Drug Discovery

Background:

  • Targeting essential enzymes in bacterial cell wall biosynthesis is crucial for developing novel antibacterial agents.
  • Pseudomonas aeruginosa, an antibiotic-resistant pathogen, possesses the essential MurF enzyme involved in cell wall precursor peptide formation.
  • MurF catalyzes peptide bond formation using UDP-MurNAc-Ala-Glu-meso-A2pm and D-Ala-D-Ala, with ATP hydrolysis.

Purpose of the Study:

  • To develop novel antibacterial agents by targeting the MurF enzyme.
  • To identify peptide ligands with high binding affinity for MurF using a phage display approach.

Main Methods:

  • Screening of a phage display 12-mer library against purified Pseudomonas aeruginosa MurF.
  • Synthesis of the MurF substrate UDP-MurNAc-Ala-Glu-meso-A2pm.
  • Development of a spectrophotometric assay to quantify MurF kinetics and inhibition.
  • Evaluation of MurFp1 inhibition kinetics and IC50/Ki values.

Main Results:

  • Identification of the MurFp1 peptide through phage display screening.
  • MurFp1 demonstrated time-dependent inhibition, becoming potent upon pre-incubation with UDP-MurNAc-Ala-Glu-meso-A2pm or ATP.
  • Inhibition was nullified by D-Ala-D-Ala during pre-incubation, indicating interference with D-Ala-D-Ala utilization.
  • MurFp1 exhibited an IC50 of 250 microM and a Ki of 420 microM (mixed type inhibition against D-Ala-D-Ala).

Conclusions:

  • MurFp1 is the first identified peptide inhibitor of the MurF enzyme.
  • The inhibitory mechanism involves interference with D-Ala-D-Ala utilization by MurF.
  • MurFp1 likely exploits UDP-MurNAc-Ala-Glu-meso-A2pm-induced structural changes for enhanced enzyme interaction.
  • MurF represents a promising target for the development of new antimicrobial drugs.

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