Purification, cloning and sequence analysis of RsrI DNA methyltransferase: lack of homology between two enzymes, RsrI and EcoRI, that methylate the same nucleotide in identical recognition sequences.
RsrI DNA methyltransferase (M-RsrI) from Rhodobacter sphaeroides has been purified to homogeneity, and its gene cloned and sequenced. This enzyme catalyzes methylation of the same central adenine residue in the duplex recognition sequence d(GAATTC) as does M-EcoRI. The reduced and denatured molecular weight of the RsrI methyltransferase (MTase) is 33,600 Da. A fragment of R. sphaeroides chromosomal DNA exhibited M.RsrI activity in E. coli and was used to sequence the rsrIM gene. The deduced amino acid sequence of M.RsrI shows partial homology to those of the type II adenine MTases HinfI and DpnA and N4-cytosine MTases BamHI and PvuII, and to the type III adenine MTases EcoP1 and EcoP15. In contrast to their corresponding isoschizomeric endonucleases, the deduced amino acid sequences of the RsrI and EcoRI MTases show very little homology. Either the EcoRI and RsrI restriction-modification systems assembled independently from closely related endonuclease and more distantly related MTase genes, or the MTase genes diverged more than their partner endonuclease genes. The rsrIM gene sequence has also been determined by Stephenson and Greene (Nucl. Acids Res. (1989) 17, this issue).
Kaszubska
W
W
Department of Biochemistry, University of Illinois, Urbana 61801.
Aiken
C
C
O'Connor
C D
CD
Gumport
R I
RI
D000595:Amino Acid Sequence; D001483:Base Sequence; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D015254:DNA Modification Methylases; D015246:Deoxyribonuclease EcoRI / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D007526:Isoelectric Point; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012045:Regulatory Sequences, Nucleic Acid; D012242:Rhodobacter sphaeroides / Q000201:enzymology*; D012689:Sequence Homology, Nucleic Acid; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity
D015254:DNA Modification Methylases; C020098:DNA modification methylase EcoRI; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific); D015246:Deoxyribonuclease EcoRI