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Turkish Journal of Biology

Subcloning and Expression of A cDNA Encoding A Schizosaccharomyces pombe Putative C-5 DNA Methyltransferase

Ergin PINARBAÞI, Hatice PINARBAÞI, David P. HORNBY

Abstract: The pmt1+ gene, which shows a striking similarity to bacterial C-5 DNA methyltransferases, has recently been isolated from Schizosaccharomyces pombe. In this study, we have subcloned pmt1+ cDNA into the bacterial expression vector pET14b, which contains a sequence coding six histidine residues upstream of the coding site so that the recombinant protein possesses a histidine tag (Hig-Tag) at its N-terminus. The constructed vector, which we have called pETSPO1, encoding the pmt1+ cDNA, was then introduced into E. coli BL21(DE3)pLysS cells and the expressed recombinant protein was purified to homogeneity by nickel-chelate-affinity chromatography. Approximately 5 mg of His-Tag-pmt1+ fusion protein was purified from one litre of induced culture.

Key Words: S. pombe, pET system, DNA Methyltransferases, protein expression, pmt1+

Turk. J. Biol., 20, (1996), 325-331.
Other articles published in the same issue: Turk. J. Biol.,vol.20,iss.4.