Process optimization for production and purification of novel fibrinolytic enzyme from Stenotrophomonas sp. KG-16-3

Abstract

Intravascular thrombosis is a major cardiovascular complication responsible for high mortality worldwide. Existing thrombolytic agents are expensive and have various side effects. As a consequence, researchers continue to search for better thrombolytic agents. Fibrinolytic proteases especially those of microbial origin are considered as potential therapeutic candidates for thrombosis. The current study reports fibrinolytic protease from a bacterial isolate Stenotrophomonas sp. KG-16-3, as it exhibits high fibrinolytic activity on fibrin agarose plate. Studies on fibrinolytic protease from Stenotrophomonas sp. are lacking. So, a detailed study was conducted for the production and purification of fibrinolytic protease. Optimizing process parameters using the Design of Experiments method enhanced the yield by 1.5-fold. The fibrinolytic enzyme was purified by ammonium sulfate precipitation, ion-exchange and gel-filtration chromatography resulting in 7.1-fold purification and 16.7% yield with specific activity of 383.8 U/mg. The purified enzyme exhibited higher fibrinolytic activity than plasmin and had a molecular weight of 39 kDa. Optimal activity of the enzyme was observed at 50 °C and pH 10. The enzyme exhibited stability up to 60 °C, over pH 7–10 and in the presence of different metal ions and solvents. The activity of the enzyme was significantly reduced in the presence of phenylmethyl sulfonyl fluoride, iodoacetic acid and 1,10-phenanthroline, suggesting that the enzyme belonged to the serine–cysteine metalloprotease category. The present study is the first ever report on the Design of Experiments based optimization of fermentation conditions for the production of fibrinolytic protease from Stenotrophomonas sp.

Keywords:

• Stenotrophomonas sp. KG-16-3
• fibrinolytic protease
• optimization
• response surface methodology
• enzyme activity
• thrombolytic agents

Acknowledgments

The author Kapila Taneja thanks Department of Science & Technology, Government of India for financial support in the form of “INSPIRE” Fellowship (IF 120559). Authors also express gratitude towards Microbial Culture Collection, NCCS (Govt. of India), Pune, for phenotypic characterization of samples.

Disclosure statement

All authors have seen the manuscript and there is no conflict of interest for the same.

Additional information

Funding

The author Kapila Taneja thanks Department of Science & Technology, Government of India for financial support in the form of “INSPIRE” Fellowship (IF 120559).