Overcoming outer membrane barrier in gram-negative bacteria by PCNP-PVP-SE1 phage endolysin: Evidence from molecular
Arman Namdari-Miraaghaie1, Mehdi Imani1, Safa Farahmand-Azar2
1Department of Basic Sciences, Faculty of Veterinary Medicine, Urmia University, Urmia, Iran.
Research in Veterinary Science
|July 28, 2026
Summary
Engineered bacteriophage endolysins show potent bactericidal activity against Gram-negative bacteria like E. coli. Fusion with PCNP enhances activity and reduces EDTA dependence, offering a promising antimicrobial strategy.
Area of Science:
- Microbiology
- Biotechnology
- Drug Discovery
Background:
- Antimicrobial resistance in Gram-negative pathogens is a growing global health threat.
- Bacteriophage endolysins are potential antimicrobials but face challenges entering Gram-negative bacteria.
- Developing strategies to overcome the outer membrane barrier is crucial for endolysin efficacy.
Purpose of the Study:
- To engineer and characterize a novel bacteriophage endolysin variant for enhanced antibacterial activity.
- To evaluate the efficacy of a PCNP-fused endolysin against Gram-negative bacteria.
- To investigate the structural and binding properties of the engineered endolysin.
Main Methods:
- Expression and purification of native and PCNP-fused endolysins.
- In vitro antibacterial assays including disk diffusion, MIC, MBC, and time-kill assays.
- In silico analyses: structural modeling, molecular docking, molecular dynamics, and MM-PBSA.
Main Results:
- The PCNP-fused endolysin exhibited significant bactericidal activity against Escherichia coli and Salmonella Typhimurium.
- Engineered variant showed larger inhibition zones and faster bacterial reduction compared to native endolysin.
- Computational analyses confirmed preserved catalytic integrity and improved binding free energy of the PCNP-fused variant.
Conclusions:
- N-terminal PCNP fusion enhances endolysin bactericidal efficacy against Gram-negative bacteria.
- The engineered endolysin demonstrates reduced dependence on EDTA for activity.
- PCNP-fused Artilysins are promising candidates for novel antimicrobial therapies.
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