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

: Premetis Georgios E, Stathi Angeliki, Papageorgiou Anastassios C, Labrou Nikolaos E

Publisher: Nature Publishing Group

: 2023

: Scientific Reports

: Scientific reports

: Sci Rep

: 7450

: 13

: 1

: 2045-2322

DOI: https://doi.org/10.1038/s41598-023-34309-2

: https://doi.org/10.1038/s41598-023-34309-2

: https://research.utu.fi/converis/portal/detail/Publication/179789465

Multidrug-resistant (MDR) bacteria have become a growing threat to public health. The gram-positive Enterococcus faecium is classified by WHO as a high-priority pathogen among the global priority list of antibiotic-resistant bacteria. Peptidoglycan-degrading enzymes (PDEs), also known as enzybiotics, are useful bactericidal agents in the fight against resistant bacteria. In this work, a genome-based screening approach of the genome of E. faecium allowed the identification of a putative PDE gene with predictive amidase activity (EfAmi1; EC 3.5.1.28) in a prophage-integrated sequence. EfAmi1 is composed by two domains: a N-terminal Zn²⁺-dependent N-acetylmuramoyl-L-alanine amidase-2 (NALAA-2) domain and a C-terminal domain with unknown structure and function. The full-length gene of EfAmi1 was cloned and expressed as a 6xHis-tagged protein in E. coli. EfAmi1 was produced as a soluble protein, purified, and its lytic and antimicrobial activities were investigated using turbidity reduction and Kirby-Bauer disk-diffusion assays against clinically isolated bacterial pathogens. The crystal structure of the N-terminal amidase-2 domain was determined using X-ray crystallography at 1.97 Å resolution. It adopts a globular fold with several α-helices surrounding a central five-stranded β-sheet. Sequence comparison revealed a cluster of conserved amino acids that defines a putative binding site for a buried zinc ion. The results of the present study suggest that EfAmi1 displays high lytic and antimicrobial activity and may represent a promising new antimicrobial in the post-antibiotic era.

s41598-023-34309-2.pdf