Open AccessCharacterization of the NgoAXP: phase‐variable type III restriction–modification system in Neisseria gonorrhoeae
Author(s) -
AdamczykPoplawska Monika,
Lower Michał,
Piekarowicz Andrzej
Publication year - 2009
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2009.01760.x
Subject(s) - neisseria gonorrhoeae , restriction enzyme , methyltransferase , biology , escherichia coli , gene , dna methyltransferase , dna , microbiology and biotechnology , methylation , restriction fragment , phase variation , dna methylation , recognition sequence , genetics , gene expression , phenotype
Methyltransferases associated with type III restriction–modification (RM) systems are phase‐variably expressed in a variety of pathogenic bacteria. NgoAXP, the type III RM system encoded by Neisseria gonorrhoeae , was characterized in this study. The cloned resngoAXP and ngoAXPmod genes were expressed in Escherichia coli strains. The restriction and modification activities of NgoAXP were confirmed in vivo by the λ phage restriction and modification test and in vitro by the methylation of DNA substrates in the presence of [ methyl ‐ 3 H]AdoMet. As in all known type III systems, the restriction activity needed the presence of both genes, while the presence of the ngoAXPmod gene was sufficient for DNA methylation. Following its overexpression, the DNA methyltransferase M.NgoAXP was purified to apparent homogeneity using metal affinity chromatography. The specific sequence recognized by this enzyme was determined as a nonpalindromic sequence: 5′‐CCACC‐3′, in which the adenine residue is methylated. We observed that in E. coli cells, the expression of the restriction phenotype associated with NgoAXP switched randomly. This phase variation was associated with the change in the number of pentanucleotide repeats (5′‐CCAAC/G‐3′) present at the 5′‐end of the coding region of the ngoAXPmod gene.