Isolation and characterization of a HpyC1I restriction-modification system in Helicobacter pylori.
Using transposon shuttle mutagenesis, we identified six Helicobacter pylori mutants from the NTUH-C1 strain that exhibited decreased adherence and cell elongation. Inverse polymerase chain reaction and DNA sequencing revealed that the same locus was interrupted in these six mutants. Nucleotide and amino acid sequences showed no homologies with H. pylori 26695 and J99 strains. This novel open reading frame contained 1617 base pairs. The amino acid sequence shared 24% identity with a putative nicking enzyme in Bacillus halodurans and 23 and 20% identity with type IIS restriction endonucleases PleI and MlyI, respectively. The purified protein, HpyC1I, showed endonuclease activity with the recognition and cleavage site 5'-CCATC(4/5)-3'. Two open reading frames were located upstream of the gene encoding HpyC1I. Together, HpyC1I and these two putative methyltransferases (M1.HpyC1I and M2.HpyC1I) function as a restriction-modification (R-M) system. The HpyC1I R-M genes were found in 9 of the 15 H. pylori strains tested. When compared with the full genome, significantly lower G + C content of HpyC1I R-M genes implied that these genes might have been acquired by horizontal gene transfer. Plasmid DNA transformation efficiencies and chromosomal DNA digestion assays demonstrated protection from HpyC1I digestion by the R-M system. In conclusion, we have identified a novel R-M system present in approximately 60% of H. pylori strains. Disruption of this R-M system results in cell elongation and susceptibility to HpyC1I digestion.
Lin
Tzu-Lung
TL
Graduate Institute of Microbiology, National Taiwan University, College of Medicine, Taipei 10016, Taiwan.
Shun
Chia-Tun
CT
Chang
Kai-Chih
KC
Wang
Jin-Town
JT
D001483:Base Sequence; D045744:Cell Line, Tumor; D017931:DNA Primers; D004262:DNA Restriction Enzymes / Q000302:isolation & purification* / Q000378:metabolism / Q000648:ultrastructure; D005816:Genetic Complementation Test; D016480:Helicobacter pylori / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D008854:Microscopy, Electron; D008969:Molecular Sequence Data
D017931:DNA Primers; D004262:DNA Restriction Enzymes