Biochemical Characterization and Application of a Detergent Stable, Antimicrobial and Antibiofilm Potential Protease from Bacillus siamensis.

Abstract

Proteases are important enzymes that are engaged in a variety of essential physiological functions and have a significant possible use in industrial applications. In this work, we reported the purification and biochemical characterization of a detergent stable, antimicrobial, and antibiofilm potential protease (SH21) produced by Bacillus siamensis CSB55 isolated from Korean fermented vegetable kimchi. SH21 was purified to obtain homogeneity via ammonium sulfate precipitation (40-80%), Sepharose CL-6B, and Sephadex G-75 column. By analyzing the SDS-PAGE and zymogram, it was determined that the molecular weight was around 25 kDa. The enzyme activity was almost completely inhibited in the presence of PMSF and DFP, which indicated that it was a member of the serine protease family. SH21 showed excellent activity with a broad range of pH and temperature, with its maximum pH of 9.0 and temperature of 55 °C. The enzyme had estimated Km and Vmax values of 0.197 mg/mL and 1.22 × 103 U/mg, respectively. In addition, it preserved good activity in the presence of different organic solvents, surfactants, and other reagents. This enzyme showed good antimicrobial activity that was evaluated by MIC against several pathogenic bacteria. Furthermore, it exhibited strong antibiofilm activity as determined by MBIC and MBEC assay and degraded the biofilms, which were analyzed by confocal microscopic study. These properties established that SH21 is a potent alkaline protease that can be used in industrial and therapeutic applications.