Premium
The Ecl 18kI restriction‐modification system: cloning, expression, properties of the purified enzymes
Author(s) -
Denjmukhametov M.M.,
Brevnov M.G.,
Zakharova M.V.,
Repyk A.V.,
Solonin A.S.,
Petrauskene O.V.,
Gromova E.S.
Publication year - 1998
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(98)00921-1
Subject(s) - restriction enzyme , tetramer , ecori , escherichia coli , dna , recognition sequence , microbiology and biotechnology , endonuclease , chemistry , restriction fragment , biochemistry , enzyme , dna methyltransferase , biology , methyltransferase , gene , methylation
Ecl 18kI is a type II restriction‐modification system isolated from Enterobacter cloaceae 18kI strain. Genes encoding Ecl 18kI methyltransferase (M. Ecl 18kI) and Ecl 18kI restriction endonuclease (R. Ecl 18kI) have been cloned and expressed in Escherichia coli . These enzymes recognize the 5′….↓CCNGG….′ sequence in DNA; M. Ecl 18kI methylates the C5 carbon atom of the inner dC residue and R. Ecl 18kI cuts DNA as shown by the arrow. The restriction endonuclease and the methyltransferase were purified from E. coli B834 [p18Ap1] cells to near homogeneity. The restriction endonuclease is present in the solution as a tetramer, while the methyltransferase is a monomer. The interactions of M. Ecl 18kI and R. Ecl 18kI with 1,2‐dideoxy‐ d ‐ribofuranose containing DNA duplexes were investigated. The target base flipping‐out mechanism is applicable in the case of M. Ecl 18kI. Correct cleavage of the abasic substrates by R. Ecl 18kI is accompanied by non‐canonical hydrolysis of the modified strand.