Improved lytic action of engineered phage-encoded endolysin and docking insights into its action on bacterial peptidoglycan.

Antimicrobial resistance in Salmonella poses significant threats, and engineered endolysins offer a promising alternative to traditional bacteriophage therapy. This study focuses on designing, developing, and evaluating a bacteriophage-derived engineered endolysin with a polycationic nonapeptide (PCNP) modification to enhance its activity. The sequence-encoded endolysin φSE218_Lys76 and its modified version φSE218_Lys76_L1_PCNP were cloned, expressed, purified, and assessed for their in-vitro antibacterial activity. Both endolysins were evaluated on live bacterial cells (with and without EDTA treatment) as well as on heat-killed bacterial cells and found antibacterial potential. The modified endolysin, φSE218_Lys76_L1_PCNP, exhibited significantly enhanced lytic activity compared to the parent endolysin, suggesting good electrostatic interactions with negatively charged bacterial surfaces. Additionally, molecular docking and dynamics simulations studies were used to check the binding affinity and structural stability of the modified endolysin on various peptidoglycan fragments. The results demonstrated a high binding affinity (-7.19 kcal/mol) and structural stability of modified endolysin with NAG-NAM tetramer, building blocks of peptidoglycan, in computational models, as compared to parent endolysin (-6.92 kcal/mol) which was found consistent with lytic activity observed in vitro. These findings highlight the potential of bacteriophage-derived endolysins as an alternative antimicrobial, offering a promising avenue for addressing the growing challenge of antibiotic-resistant pathogens.