Apoptin NLS2 homodimerization strategy for improved antibacterial activity and bio-stability

Area of Science:

• Biochemistry
• Microbiology
• Drug Discovery

Background:

• Antibiotic resistance necessitates novel antimicrobial agents.
• Antimicrobial peptides (AMPs) are a promising alternative.
• Apoptin nuclear localization sequence (NLS2)-derived peptides offer potential.

Purpose of the Study:

• To investigate the antimicrobial activity of an NLS2-derived peptide (ANLP).
• To evaluate the efficacy of NLS dimerization for enhancing AMP properties.
• To assess the bio-stability and safety of dimeric AMPs.

Main Methods:

• Peptide synthesis of ANLP and its homodimer variants (ANLP-K1, ANLP-K2).
• Antimicrobial activity testing against *Escherichia coli* and *Salmonella*.
• Serum stability assays, cytotoxicity tests on Vero cells, dye penetration assays, zeta potential analysis, and scanning electron microscopy.

Main Results:

• ANLP-K2 demonstrated potent antibacterial activity (12 µM MIC).
• Dimeric peptides showed significantly improved bio-stability compared to linear ANLP.
• Dimeric AMPs exhibited membrane interaction, effective charge neutralization of LPS and bacterial cells, and induced pore formation without cytotoxicity.

Conclusions:

• Lysine scaffold dimerization of Apoptin NLS2 enhances antibacterial efficacy and bio-stability.
• This strategy effectively neutralizes LPS and offers a template for novel AMP design.
• NLS-based dimeric peptides represent a promising avenue for combating antibiotic resistance.