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

• Published in: Amino acids Vol. 55; no. 10; pp. 1405 - 1416
• Main Authors: Kumari, Anu, Singh, Mahavir, Sharma, Ruchi, Kumar, Tarun, Jindal, Naresh, Maan, Sushila, Joshi, Vinay G.
• Format: Journal Article
• Language: English
• Published: Vienna Springer Vienna 01.10.2023; Springer Nature B.V
• Subjects: Analytical Chemistry; Antibacterial activity; Antibiotic resistance; Antimicrobial activity; Antimicrobial agents; Antimicrobial peptides; Biochemical Engineering; Biochemistry; Biomedical and Life Sciences; Cytotoxicity; Dimerization; Dimers; Dye penetrants; E coli; Life Sciences; Lipopolysaccharides; Localization; Lysine; Minimum inhibitory concentration; Neurobiology; Original Article; Peptides; Proteomics; Scanning electron microscopy; Stability; Vero cells; Zeta potential; Peptide homodimer; Antibacterial activity; Apoptin; Nuclear localization sequence; Lipopolysaccharides
• ISSN: 0939-4451; 1438-2199
• DOI: 10.1007/s00726-023-03321-1

The emergence of antibiotic resistance prompts exploration of viable antimicrobial peptides (AMPs) designs. The present study explores the antimicrobial prospects of Apoptin nuclear localization sequence (NLS2)-derived peptide ANLP (PRPRTAKRRIRL). Further, we examined the utility of the NLS dimerization strategy for improvement in antimicrobial activity and sustained bio-stability of AMPs. Initially, the antimicrobial potential of ANLP using antimicrobial peptide databases was analyzed. Then, ANLP along with its two homodimer variants namely ANLP-K1 and ANLP-K2 were synthesized and evaluated for antimicrobial activity against Escherichia coli and Salmonella . Among three AMPs, ANLP-K2 showed efficient antibacterial activity with 12 µM minimum inhibitory concentration (MIC). Slow degradation of ANLP-K1 (26.48%) and ANLP-K2 (13.21%) compared with linear ANLP (52.33%) at 480 min in serum stability assay indicates improved bio-stability of dimeric peptides. The AMPs presented no cytotoxicity in Vero cells. Dye penetration assays confirmed the membrane interacting nature of AMPs. The zeta potential analysis reveals effective charge neutralization of both lipopolysaccharide (LPS) and bacterial cells by dimeric AMPs. The dimeric AMPs on scanning electron microscopy studies showed multiple pore formations on the bacterial surface. Collectively, proposed Lysine scaffold dimerization of Apoptin NLS2 strategy resulted in enhancing antibacterial activity, bio-stability, and could be effective in neutralizing the off-target effect of LPS. In conclusion, these results suggest that nuclear localization sequence with a modified dimeric approach could represent a rich source of template for designing future antimicrobial peptides.