CLONING AND EXPRESSION OF TRUNCATED AND INTACT STREPTOKINASE MOLECULES IN E.COLI AND EVALUATION OF THEIR BIOLOGICAL ACTIVITIES

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ARABI R.; ROOHVAND F.; NOROUZIAN D.; AGHASADEGHI M.R.; MEMARNEZHADIAN A.; KHANAHMAD H.; SADAT M.

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Journal: JOURNAL OF AGRICULTURAL BIOTECHNOLOGY Year:2010 | Volume:2 | Issue:1 Page(s): 55-67

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Title

CLONING AND EXPRESSION OF TRUNCATED AND INTACT STREPTOKINASE MOLECULES IN E.COLI AND EVALUATION OF THEIR BIOLOGICAL ACTIVITIES

Author(s)

Keywords

STREPTOKINASEQ3

TRUNCATED PROTEINSQ3

CLONINGQ2

BIOLOGICAL ACTIVITYQ3

Abstract

STREPTOKINASE is the most utilized drug for thrombolytic therapy. However, due to its nonhuman origin and nonspecific activity in the absence of fibrin, its application may encounter side effects such as allergic reactions and hemorrhage respectively. It is suggested that some truncated forms of STREPTOKINASE may show better fibrin-specificity and decreased antigenicity while retaining most of the enzymatic activity. In the present study, different truncated forms of STREPTOKINASE gene, “Skc60-386 and skc143-386” together with the intact form of skc (skc1-414) were cloned in T5 and T7 based-E.coli expression vectors, “pQE30and pET41a” respectively. While expression of intact STREPTOKINASE could equally be achieved in both systems, but pET41a was the only effective system to produce truncated forms. pET41a vector contains three tandem tags before multiple CLONING sites which possibly prevent intrinsic problems encountered in heterologous recombinant protein expression in E.coli. Analyses of the purified proteins by SDS-PAGE and Western blotting evidenced for the true expression and the expected MWs. While caseinolysis method was notable to demonstrate exact difference between specific activity of SK143-386, SK60-386 and SK1-414 but truncated molecules showed high reduction of specific activity (83 to 91percent) compared to the full-length STREPTOKINASE by more exact techniques such as colorimetric assay. This study provided a primary comparative study for expression of STREPTOKINASE and its truncated forms and analyses of their specific activity. Thus further analyses would determine their specificity to fibrin and antigenicity.

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APA: Copy

ARABI, R., ROOHVAND, F., NOROUZIAN, D., AGHASADEGHI, M.R., MEMARNEZHADIAN, A., KHANAHMAD, H., & SADAT, M.. (2010). CLONING AND EXPRESSION OF TRUNCATED AND INTACT STREPTOKINASE MOLECULES IN E.COLI AND EVALUATION OF THEIR BIOLOGICAL ACTIVITIES. JOURNAL OF AGRICULTURAL BIOTECHNOLOGY, 2(1), 55-67. SID. https://sid.ir/paper/224395/en

Vancouver: Copy

ARABI R., ROOHVAND F., NOROUZIAN D., AGHASADEGHI M.R., MEMARNEZHADIAN A., KHANAHMAD H., SADAT M.. CLONING AND EXPRESSION OF TRUNCATED AND INTACT STREPTOKINASE MOLECULES IN E.COLI AND EVALUATION OF THEIR BIOLOGICAL ACTIVITIES. JOURNAL OF AGRICULTURAL BIOTECHNOLOGY[Internet]. 2010;2(1):55-67. Available from: https://sid.ir/paper/224395/en

IEEE: Copy

R. ARABI, F. ROOHVAND, D. NOROUZIAN, M.R. AGHASADEGHI, A. MEMARNEZHADIAN, H. KHANAHMAD, and M. SADAT, “CLONING AND EXPRESSION OF TRUNCATED AND INTACT STREPTOKINASE MOLECULES IN E.COLI AND EVALUATION OF THEIR BIOLOGICAL ACTIVITIES,” JOURNAL OF AGRICULTURAL BIOTECHNOLOGY, vol. 2, no. 1, pp. 55–67, 2010, [Online]. Available: https://sid.ir/paper/224395/en

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