Induced expression of <Emphasis Type="Italic">Eco</Emphasis>RI endonuclease as an active maltose-binding fusion protein in <Emphasis Type="Italic">Escherichia coli</Emphasis> |
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Authors: | A Gholizadeh M H Faizi B Baghban Kohnehrouz |
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Institution: | 1.Research Institute for Fundamental Sciences (RIFS),University of Tabriz,Tabriz,Iran;2.Department of Biology,University of Tabriz,Tabriz,Iran |
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Abstract: | Among the numerous bacterial Type II restriction enzymes, EcoRI endonuclease is the most extensively studied and is widely used in recombinant DNA technology. Its heterologous overexpression
as recombinant protein has already been studied. However, very limited information concerning its fused product is available
thus far. In the present study, the EcoRI restriction endonuclease gene was cloned and expressed as a part of maltose-binding fusion protein under the control of
strong inducible tac promoter in TB1 strain of Escherichia coli cells. Transformed cells containing pMALc2X-EcoRI recombinant plasmid were unable to grow under experimental conditions. However, fused EcoRI protein was purified (with the yield of 0.01 mg/l of bacterial culture) by affinity chromatography from E. coli cells induced at the late exponential phase of growth. Restriction quality test revealed that the purified product could
restrict a control plasmid DNA in vitro. |
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