<i>In Vitro</i> Antimycobacterial Activities of Short Peptide-Functionalized Silver Nanoparticle and Its <i>In Silico</i> Mechanistic Insight

The rise of drug-resistant microbes has made antimicrobial therapy increasingly challenging, and despite several reports on peptide-functionalized silver nanoparticles, their efficacy against <i>Mycobacterium</i> species remains largely unexplored. In this study, we synthesized short peptide functionalized silver nanoparticles to develop an effective antimycobacterial agent, where peptides acted as both reducing and stabilizing agents for the one-pot synthesis of silver nanoparticles (AgNPs). The developed nanoparticles were characterized by high-resolution transmission electron microscopy (HR-TEM), dynamic light scattering (DLS), Fourier transform infrared spectroscopy (FTIR), and UV-visible spectroscopy (UV-vis). The positively charged peptide-capped silver nanoparticles exhibited significant antimycobacterial activity against acid-fast mycobacterial strains, including <i>Mycobacterium smegmatis</i>, <i>Mycobacterium bovis</i>, and <i>Mycobacterium marinum</i>, compared to peptides alone, which could be due to the integrated effect of the peptide-functionalized AgNPs. Among the synthesized nanoparticles, linear peptide 2 (LP 2) functionalized AgNP exhibited the highest antimycobacterial efficacy against the <i>Mycobacterium</i> strains, with the lowest MIC (5 μM). AgNP LP 2 was found to be efficient to penetrate the mycobacterial cell wall, inducing membranolytic activity, triggering oxidative stress and degrading DNA, which led to the death of mycobacterial cells. Molecular docking and molecular dynamics (MD) simulations of the peptides with key enzyme FadD32 (<i>Msm</i>FadD32), <i>Mycobacterium smegmatis</i>, demonstrated strong interactions near the active site cleft, indicating potential inhibition of the mycolic acid biosynthesis pathway by the LP 2 peptide. This disruption likely challenges the organism's pathogenicity and supports the peptides' role in contributing to membranolytic activity. Additionally, AgNP LP 2 demonstrated the ability to inhibit biofilm formation and effectively disrupt preformed mycobacterial biofilms while exhibiting negligible cytotoxicity toward human embryonic kidney (HEK293) cells. In summary, our results suggest that newly developed AgNPs exhibit antimycobacterial activity without compromising the cell viability of normal cells, making them highly potent as prospective antimycobacterial agents.