Cloning and expression of hybrid streptokinase towards clot-specific activity.
Streptokinase (SK) is a thrombolytic agent that is widely used to treat myocardial infarction and pulmonary embolism. The lack of fibrin specificity of SK for the clot lysis is one of the limitations of SK. In this study, we have incorporated the finger and Kringle 2 domains from the human tissue type plasminogen activator gene (t-PA) at the 5' end of the SK gene. These domains are responsible for specific binding to fibrin. We have used the pRSETB vector in an attempt to express the hybrid streptokinase possessing specificity for fibrin. On this regard, three hybrid streptokinase were constructed and expressed in Escherichia coli BL21 (DE3): the finger domain with SK (FSK), the Kringle 2 domain with SK (KSK) and the finger domain+Kringle 2 with SK (FKSK). The activities of the hybrid SKs were assessed by caseinolytic assay and clot lysis assay. All hybrid SKs were found to activate plasminogen in the caseinolytic plate assay. In the clot lysis assay, KSK and FSK were able to dissolute human blood and artificial clots in a fibrin-dependent manner unlike the SK and FKSK proteins.
Department of Biology, College of Education for Pure Science, University of Anbar, Ramadi, Iraq. Electronic address: harithkamil@yahoo.com.
Harith K
HK
Buniya
Genetic Engineering Unit, Center for Biotechnology, Anna University, Guindy, Chennai- 600 025, India.
Vadivel
V
Murugan
Genetic Engineering Unit, Center for Biotechnology, Anna University, Guindy, Chennai- 600 025, India.
Chinnathambi
C
Thangadurai
D003001:Cloning, Molecular / Q000379:methods; D005337:Fibrin / Q000235:genetics; D063285:Fibrin Clot Lysis Time / Q000379:methods; D006801:Humans; D018082:Kringles / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics*; D013300:Streptokinase / Q000235:genetics*; D010959:Tissue Plasminogen Activator / Q000235:genetics
D011994:Recombinant Proteins; D005337:Fibrin; D013300:Streptokinase; D010959:Tissue Plasminogen Activator
Escherichia coli; Hybrid streptokinase; Plasminogen activator; Thrombolytic agents; t-PA