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Abstract To obtain the fusion protein of Lycopersicon esculentum Metacaspase (LeM) and study the characteristics and function of it, we cloned the open reading frame of LeM in Lycopersicon esculentum by RT-PCR, and successfully constructed the prokaryotic expression vector pET28a-LeM for LeM expression. The recombinant plasmid pET28a-LeM was transformed into Escherichia coli BL21(DE3)PlysS and Rosetta for expression, induced by IPTG, and the conditions of the expression was optimized. SDS-PAGE revealed that the best expression was induced by 1 mmol/L IPTG at 37℃ and the recombinant protein expressed mainly as inclusion body in E. coli BL21(DE3)PlysS while as fusion protein in E. coli Rosetta at the same condition. Enzyme activity assay showed that the activity of LeM in E.coli Rosetta was 33 RFU/min, which was much higher than 7 RFU/min of E. coli BL21(DE3)PlysS. We demonstrate that the strain has a great influence on the expression of LeM fusion protein.
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Received: 12 July 2010
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