Abstract
Streptokinase (SK) is one of the essential fibrinolytic agents and is commonly employed to treat myocardial infarction or heart attack. Despite its significance as an anti-clot agent, SK exhibits certain drawbacks, including limited half-life in the bloodstream, non-specific to fibrin, and may cause process workers to develop undesirable immune responses. Acknowledging the high specificity of staphylokinase (SAK) towards fibrin and the powerful recombinant DNA technology, this study was carried out to evaluate the fibrinolytic activity of a recombinant staphylokinase-streptokinase (SAK-SK) protein in Escherichia coli BL21(DE3). Briefly, the SAK gene isolated from Staphylococcus aureus was ligated at the 5’end of the SK gene using the pGEM®-3Zf (+) cloning vector and transformed into the expression host E. coliBL21(DE3) before being induced with isopropyl β-D-1-thiogalactopyranoside (IPTG).Subsequently, the produced recombinant SK protein was analyzed using 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).The biological performance of the recombinant SK protein was then examined via caseinolytic and fibrinolytic assays and compared to that of the native SK protein. Based on the results, the recombinant SK protein contained 563 amino acid residues with a molecular weight of approximately 63.1 kDa. The hybrid SK protein was more active in the casein lysis experiment compared to the native SK, with a more significant halo zone formation. Moreover, the hybrid SK protein demonstrated a complete clot lysis compared to the poor clot lysis of the native SK protein. In conclusion, this study successfully developed a recombinant hybrid SK protein with enhanced affinity to fibrin, making it a potential anti-clot agent.
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Abbreviations
- E. coli :
-
Escherichia coli
- F:
-
Forward
- R:
-
Reverse
- PCR:
-
Polymerase Chain Reaction
- Rpm:
-
Revaluation per minute
- SK:
-
Streptokinase
- SAK:
-
Staphylokinase
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Acknowledgements
For their assistance in completing this work, the authors are grateful to all of the employees at the Department of Biology, College of Education for Pure Science, University Of Anbar, as well as their colleagues at the Molecular Biology Lab.
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Buniya, H.K., Hameed, A.K. & Al-Hayawi, A.Y. Cloning and expression of staphylokinase-streptokinase recombinant protein in E. coli BL21(DE3). Biologia 78, 1113–1117 (2023). https://doi.org/10.1007/s11756-023-01311-y
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DOI: https://doi.org/10.1007/s11756-023-01311-y