Detergent-Stable Alkaline Protease from Marine <i>Bacillus subtilis</i> BGN4: Production, Characterization, and Anti-Biofilm Activity

The study presents the alkaline protease production, purification, and enzyme characterization of a halo-stable protease from a marine bacterium, Bacillus subtilis BGN4 (MN093306). Profiling studies revealed increased bacterial growth and enzyme production following 120 hours of incubation at a temperature of 45°C, a pH of 11.0, and a sodium chloride at 5% (w/v). Fructose at 1% (w/v) and tryptone at 0.5% (w/v) were used as the preferred carbon and nitrogen sources, respectively. The process of purifying the enzyme involved DEAE-cellulose ion-exchange chromatography and Sephadex G-75 column chromatography. The enzyme characterization demonstrated that the protease had a molecular mass of 32 kDa, as determined using SDS-PAGE and casein zymography. Studies showed that the enzyme maintained a higher activity across a broad spectrum of temperatures (20–80°C), NaCl (0–5%), and pH (6–12). Protease exhibited the highest stability and activity at 40°C, 3% NaCl, and pH 10.0. The kinetic enzyme studies showed the K m and V max values as 0.054 mg/mL and 17.85 U/mL, respectively. Halo-stable alkaline protease exhibited better compatibility with numerous commercial detergent formulations. The enzyme had a potential biofilm activity (&gt;85%) against Staphylococcus aureus NCIM 2654 and Pseudomonas aeruginosa NCIM 5514. Detergent compatibility and antibiofilm action of the protease produced by B. subtilis BGN4 find economical and sustainable applications in agriculture, health care, and industry.