Purification and Characterisation of a Fibrinolytic Enzyme from Rhizopus microsporus var. tuberosus.

Nan Zhang,Shuli Zhang,Yingdong Wang,Xingnan Cao,Zhe Sun,Haikuan Wang,Yan Shi

doi:10.17113/ftb.53.02.15.3874

Nan Zhang, Shuli Zhang + Show 5 more

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https://doi.org/10.17113/ftb.53.02.15.3874

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Abstract

Extracellular fibrinolytic enzyme from Rhizopus microsporus var. tuberosus was purified and characterised. The microorganism was isolated in a distillery from daqu, a fermentative agent used in the production of Chinese liquor and vinegar at different temperatures. The fibrinolytic enzyme was partially purified by ammonium sulphate precipitation, dialysis, DEAE Sepharose® Fast Flow ion exchange chromatography and Sephadex G-75 gel ﬁltration chromatography. The molecular mass of the fibrinolytic enzyme was estimated to be 24.5 kDa by SDS-PAGE. The purified enzyme showed optimal activity at pH=7.0 and 37 °C by fibrin plate method. It showed stronger resistance to the inhibition by trypsin and was stable at 37 °C retaining 96.1% residual activity after 4 h of incubation. The fibrinolytic activity of the enzyme was enhanced by Na+, Ca2+, Mg2+ and Mn2+. Conversely, Zn2+ and Cu2+ partly inhibited enzymatic activity. Using fibrin plate method, we found that the enzyme not only degrades fibrin directly, but also activates plasminogen into plasmin to degrade fibrin. The results indicate that the pure enzyme has a potential in dissolving blood clot, and the possibility for application in the treatment of thrombosis.

Highlights

SummaryExtracellular fibrinolytic enzyme from Rhizopus microsporus var. tuberosus was purified and characterised
Cardiovascular diseases, such as acute myocardial infarction, ischaemic heart disease and high blood pressure, are the leading causes of death in the world [1]
The results indicate that the pure enzyme has a potential in dissolving blood clot, and the possibility for application in the treatment of thrombosis

Summary

Extracellular fibrinolytic enzyme from Rhizopus microsporus var. tuberosus was purified and characterised. Extracellular fibrinolytic enzyme from Rhizopus microsporus var. The fibrinolytic enzyme was partially purified by ammonium sulphate precipitation, dialysis, DEAE Sepharose® Fast Flow ion exchange chromatography and Sephadex G-75 gel filtration chromatography. The molecular mass of the fibrinolytic enzyme was estimated to be 24.5 kDa by SDS-PAGE. The purified enzyme showed optimal activity at pH=7.0 and 37 °C by fibrin plate method. It showed stronger resistance to the inhibition by trypsin and was stable at 37 °C retaining 96.1 % residual activity after 4 h of incubation. The fibrinolytic activity of the enzyme was enhanced by Na+, Ca2+, Mg2+ and Mn2+. We found that the enzyme degrades fibrin directly, and activates plasminogen into plasmin to degrade fibrin.

Introduction

Materials and Methods

Results and Discussion

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