Nucleotide sequence of a novel kanamycin resistance gene, aphA-7, from Campylobacter jejuni and comparison to other kanamycin phosphotransferase genes.
A novel kanamycin phosphotransferase gene, aphA-7, was cloned from a 14-kb plasmid obtained from a strain of Campylobacter jejuni and the nucleotide sequence of the gene was determined. The presumed open reading frame of the aphA-7 structural gene was 753 bp in length and encoded a protein of 251 amino acids with a calculated weight of 29,691 Da. A 29-kDa protein was demonstrated in Escherichia coli maxicells containing the cloned aphA-7 gene. A ribosomal binding site corresponding to 5 of 8 bases of the 3' end of the E. coli 16S rRNA was 8 bp upstream of the start codon. Sequences corresponding to the -35 and -10 regions of the consensus promoter sequences of E. coli were upstream of the presumed initiation codon of the gene. The DNA sequence was most closely related to the aphA-3 gene from Streptococcus faecalis, showing 55.4% sequence similarity. There was 45.6% identity at the amino acid level between the aphA-3 and the aphA-7 proteins. Of the three conserved regions noted previously in phosphotransferase genes, the aphA-7 amino acid sequence was identical to the six conserved amino acids in motif 3, but differed in one of the five conserved amino acids in motif 1 (if gaps are permitted) and 3 of the 10 conserved residues in motif 2. The 32.8% G + C ratio in the open reading frame of the aphA-7 kanamycin resistance gene, which is similar to that of the C. jejuni chromosome, suggests that the aphA-7 may be indigenous to Campylobacters.
Tenover
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Seattle Veterans Administration Medical Center, Washington 98108.
Gilbert
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O'Hara
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D000595:Amino Acid Sequence; D000617:Aminoglycosides / Q000494:pharmacology; D001483:Base Sequence; D002168:Campylobacter fetus / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D002678:Chimera; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D004352:Drug Resistance, Microbial; D004591:Electrophoresis, Polyacrylamide Gel; D005798:Genes, Bacterial; D007612:Kanamycin / Q000235:genetics*; D007613:Kanamycin Resistance / Q000235:genetics*; D008969:Molecular Sequence Data; D010770:Phosphotransferases / Q000235:genetics*; D010957:Plasmids; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid
D000617:Aminoglycosides; D004269:DNA, Bacterial; D007612:Kanamycin; D010770:Phosphotransferases