In silico characterization of an engineered Artilysin^®: structural insights into PCNP-fused PVP-SE۱ endolysin and peptidoglycan interactions

The rise of antimicrobial resistance poses a severe global health challenge, with multidrug-resistant bacteria causing millions of deaths annually and diminishing the effectiveness of traditional antibiotics. Bacteriophage-derived endolysins, such as PVP-SE۱gp۱۴۶ from Salmonella phage PVP-SE۱, emerge as promising alternatives due to their high specificity, low toxicity, and minimal resistance development. However, their activity against Gram-negative bacteria is limited by poor outer membrane (OM) penetration. This study investigates the structural and functional impacts of fusing the polycationic nonapeptide, PCNP to PVP-SE۱gp۱۴۶ to create engineered endolysin that enhance OM permeability while preserving enzymatic function. Using advanced computational tools including Phyre۲, GalaxyWEB, and AlphaFold۲, three-dimensional models of native and engineered endolysins were generated and validated with Verify۳D. Sequence analysis via Basic Local Alignment Search Tool identified homologs, and structural superposition with template modelin-align (TM-align) confirmed high conservation (TM-score ۰.۹۲۱, Root Mean Square Deviation ۱.۸۴ Å) despite the PCNP insertion. Active sites were predicted using PrankWeb, and molecular docking with AutoDock assessed interactions with peptidoglycan components (e.g., N-acetylmuramic acid - N-acetylglucosamine dimer, tetramer). Results revealed superior binding affinities in the native endolysin (e.g., – ۱۰.۳۵ kcal mol-۱ for N-acetylmuramic acid -L-alanine) compared to the engineered variant (– ۹.۶۵ kcal mol-۱ for N-acetylmuramic acid - N-acetylglucosamine -L-alanine trimer), with positive energies for larger ligands indicating steric hindrance. This trade-off reduced affinity for improved OM penetration supports potential enhancement of OM penetration while preserving bactericidal function like Pseudomonas aeruginosa. The findings highlight the potential of rational engineering for superior antimicrobials, emphasizing the need for molecular dynamics simulations and experimental validation to optimize Artilysin® design in combating antimicrobial resistance.

کلیدواژه ها:

antimicrobial resistance ، Artilysin&reg ، Endolysin modeling ، Molecular Docking ، Peptidoglycan interactions