Clot Buster Enzyme Activity from Bacillus sp.,

Abstract:

Context: Clot buster enzyme is widely used as the thrombolytic agent due to its low cost as compared to several other thrombolytic agents. Aim: The aim of the study is to isolate clot buster enzyme producing bacteria from soil sample. Materials and Methods: In the present study, we have isolated 20 different bacterial strains from soil source collected near meat shops and screened for the production of clot buster enzymes. The sample was serially diluted and characterized in the Bacillus differential medium. Most promising clot buster enzymes producing bacterial strain were selected by the initial screening of the casein hydrolase assay. The one strain showed activity with a zone of clearance in the casein hydrolase assay were characterized by various morphological and biochemical test. Morphological identification of the bacterial cells was examined by Gram staining and by SEM analysis. Production, media was prepared and inoculated by potent strain VITSM04 and processed for seven days of fermentation. The enzyme was purified by 70% ammonium sulphate precipitation, 50kDa ultra filtration and gel filtration by Sephadex G -100. Results: The crude enzyme was found to show strong clot buster enzyme activity with specific activity of 640.80U/mg in clot lysis of human blood. The concentration of the protein was found to be 55.12mg/ml. The molecular weight of clot buster enzyme was detected by 10% SDS-PAGE in single band with low molecular weight of 28kDa. Clot buster enzyme was quantified using an HPLC system with retention time of 4 min by the refractive index detector. Conclusion: Thus the enzyme can be used in the release of blood clot and fibrinolysis disorder which is a serious medical problem. The successful development of potent strain VITMS07 can holds the promising clot buster enzyme activity can be used in industrial application with large scale production.

Key words: Cardiovascular, Clot buster, Casein hydrolysis, SDS-PAGE, HPLC.