Optimization of subtilisin production from Bacillus subtilis strain ZK3 and biological and molecular characterization of synthesized subtilisin capped nanoparticles.

ABSTRACT

The increase and dissemination of multi-drug resistant bacteria have presented a major healthcare challenge, making bacterial infections a significant concern. The present research contributes towards the production of bioactive subtilisin from a marine soil isolate Bacillus subtilis strain ZK3. Custard apple seed powder (raw carbon) and mustard oil cake (raw nitrogen) sources showed a pronounced effect on subtilisin production. A 7.67-fold enhancement in the production was evidenced after optimization with central composite design-response surface methodology. Subtilisin capped silver (AgNP) and zinc oxide (ZnONP) nanoparticles were synthesized and characterized by UV-Visible spectroscopy. Subtilisin and its respective nanoparticles revealed significant biological properties such as, antibacterial activity against all tested pathogenic strains with potential against Escherichia coli and Pseudomonas aeruginosa. Prospective antioxidant behavior of subtilisin, AgNP and ZnONP was evidenced through radical scavenging assays with ABTS and DPPH. Subtilisin, AgNP and ZnONP revealed cytotoxic effect against cancerous breast cell lines MCF-7 with IC50of 83.48, 3.62 and 7.57 µg/mL respectively. Characterizations of nanoparticles were carried out by Fourier transform infrared spectroscopy, scanning electron microscopy with energy dispersive X-ray, X-ray diffraction, thermogravimetric analysis and atomic force microscopy analysis to elucidate the structure, surface and thermostability properties. The study proposes the potential therapeutic applications of subtilisin and its nanoparticles, a way forward for further exploration in the field of healthcare.