[Gene cloning, comparative analysis of the protein structures from Fsp4HI restriction-modification system and biochemical characterization of the recombinant DNA methyltransferase]

Abstract

Genes coding for the restriction-modification system Fsp4HI, recognizing the sequence 5'-GCNGC-3' have been cloned in Escherichia coli ER2267 cells and its primary structure has been determined. This RM system consists of two genes: the DNA-methyltransferase gene which is followed by the restriction endonuclease gene in the same direction. The analysis of amino acid sequences of the proteins showed that M.Fsp4HI belongs to C5 DNA-methyltransferases, and the restriction enzyme shares more or less significant homology to just a few restriction endonucleases with related recognition sequences. M.Fsp4HI enzyme was purified by means of column chromatography. According to the results of biochemical study it was considered that M.Fsp4HI has its optimal activity at 30 degree C and pH 7.5. M.Fsp4HI modifies the first cytosine residue in the sequence 5'-GCNGC-3'.