In vivo specificity of EcoRI DNA methyltransferase.
The EcoRI adenine DNA methyltransferase forms part of a bacterial restriction/modification system; the methyltransferase modifies the second adenine within the canonical site GAATTC, thereby preventing the EcoRI endonuclease from cleaving this site. We show that five noncanonical EcoRI sites (TAATTC, CAATTC, GTATTC, GGATTC and GAGTTC) are not methylated in vivo under conditions when the canonical site is methylated. Only when the methyltransferase is overexpressed is partial in vivo methylation of the five sites detected. Our results suggest that the methyltransferase does not protect host DNA against potential endonuclease-mediated cleavage at noncanonical sites. Our related in vitro analysis of the methyltransferase reveals a low level of sequence-discrimination. We propose that the high in vivo specificity may be due to the active removal of methylated sequences by DNA repair enzymes (J. Bacteriology (1987), 169 3243-3250).
Smith
D W
DW
Department of Chemistry, University of California, Santa Barbara 93106.
Crowder
S W
SW
Reich
N O
NO
D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D008745:Methylation; D010957:Plasmids; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000378:metabolism*; D013379:Substrate Specificity
D004247:DNA; C020098:DNA modification methylase EcoRI; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific)