Baculovirus-mediated high level expression of a mammalian DNA methyltransferase.
The murine C-5 cytosine DNA methyltransferase (MTase, E.C.2.1.1.37) containing a hexahistidine affinity leader peptide has been expressed at levels which are at least 50-fold higher than previously reported. The recombinant enzyme has activity levels similar to the wild-type enzyme. The recombinant polypeptide binds to and elutes from a nickel affinity resin (IMAC resin). No dramatic differences in post-translational modification between the wild-type and recombinant enzyme were observed. The recombinant system will be useful in performing site-directed mutagenesis and will facilitate enzymological and biological investigations of this enzyme.
Glickman
J F
JF
Department of Chemistry, University of California, Santa Barbara 93106.
Reich
N O
NO
D000818:Animals; D016367:Baculoviridae; D015153:Blotting, Western; D002460:Cell Line; D002846:Chromatography, Affinity; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D051379:Mice; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D018411:Spodoptera; D014162:Transfection / Q000379:methods
D011994:Recombinant Proteins; D004248:DNA (Cytosine-5-)-Methyltransferases