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Structural and functional features of a broad-spectrum prophage-encoded enzybiotic from Enterococcus faecium

Georgios E Premetis1, Angeliki Stathi2, Anastassios C Papageorgiou3

  • 1Laboratory of Enzyme Technology, Department of Biotechnology, School of Applied Biology and Biotechnology, Agricultural University of Athens, 75 Iera Odos Street, 11855, Athens, Greece.

Scientific Reports
|May 8, 2023
PubMed

Insights

Researchers identified a novel enzyme, EfAmi1, from Enterococcus faecium with potent antimicrobial activity. This peptidoglycan-degrading enzyme shows promise as a new therapeutic agent against multidrug-resistant bacteria in the post-antibiotic era.

Area of Science:

  • Microbiology
  • Biochemistry
  • Structural Biology

Background:

  • Multidrug-resistant (MDR) bacteria, particularly Gram-positive Enterococcus faecium, pose a significant global public health threat.
  • Peptidoglycan-degrading enzymes (PDEs), or enzybiotics, are emerging as crucial bactericidal agents against antibiotic-resistant pathogens.
  • E. faecium is a WHO-classified high-priority pathogen, necessitating novel therapeutic strategies.

Purpose of the Study:

  • To identify and characterize novel peptidoglycan-degrading enzymes (PDEs) from E. faecium with antimicrobial potential.
  • To investigate the lytic and antimicrobial activities of the identified enzyme, EfAmi1.
  • To determine the crystal structure of the N-terminal domain of EfAmi1 for mechanistic insights.

Main Methods:

  • Genome-based screening of E. faecium to identify putative PDE genes.
  • Cloning, expression, and purification of the full-length EfAmi1 protein in E. coli.
  • Turbidity reduction and Kirby-Bauer disk-diffusion assays to evaluate lytic and antimicrobial activity.
  • X-ray crystallography to determine the structure of the N-terminal amidase-2 domain.

Main Results:

  • Identification of a novel putative PDE gene, EfAmi1, encoding a Zn2+-dependent N-acetylmuramoyl-L-alanine amidase-2 (NALAA-2) domain.
  • EfAmi1 was successfully expressed as a soluble, purified 6xHis-tagged protein.
  • Demonstrated high lytic and antimicrobial activity against clinically relevant bacterial pathogens.
  • The crystal structure of the N-terminal domain revealed a globular fold with a putative zinc-binding site.

Conclusions:

  • EfAmi1 exhibits significant lytic and antimicrobial properties, highlighting its potential as a novel therapeutic agent.
  • The structural characterization provides insights into the enzyme's mechanism of action.
  • EfAmi1 represents a promising candidate for combating infections caused by multidrug-resistant bacteria in the post-antibiotic era.

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