PURIFICATION AND CHARACTERIZATION OF EXTRACELLULAR PROTEASE FROM SOIL ISOLATE Stenotrophomonas maltophilia AS NOVEL TARGET FOR FIBRINOLYSIS

Abstract

Fibrinolytic protease has a potential role as thrombolytic agent and useful in cardiovascular disease (CVD) treatment. In this study, a potent fibrinolytic protease-producing bacterium was isolated from casein growth medium and identified as Stenotrophomonas maltophilia by characterizing biochemical tests and 16 s rRNA sequencing. The protease production was carried out by submerged fermentation and further purified by ammonium sulphate precipitation, dialysis and ion-exchange chromatography methods. The specific activity of protease significantly increased with step by step of purification process and finally became 1.87 U/mg protein with a purification fold of 1.68 and yield of 59.52%. The sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed molecular weight of purified protease as ~47kDa, respectively. To the authors’ knowledge, this is the first report of ∼47 kDa protease from the bacterial strain Stenotrophomonas maltophilia. The characterized enzyme exhibits maximal enzyme activity at pH 8 and temperature 40°C. The activity of enzyme is activated by cations Na+, K+, Ca++ and Mg++, whereas significant loss of activity was observed with EDTA, Zn++, Cu++ and Hg++. The Lineweaver-Burk plot analysis showed a km value of 0.303 mg/ml and Vmax as 0.00714. The present study indicates that fibrinolytic protease produced from the Stenotrophomonas maltophilia KJ801664 has an antioxidant property by 1,1-diphenyl-2-picryl-hydrazyl (DPPH) method.