Molecular characterization of the gyrB region of the oral spirochete, Treponema denticola.
The nucleotide (nt) sequence of the Treponema denticola (Td) DNA gyrase beta-subunit gene (gyrB) has been determined. Southern blot analysis of Td chromosomal DNA indicated that gyrB is present as a single copy. Approximately 3.2kb of the nt sequence 5' and 0.7kb of nucleotide sequence 3' of gyrB were obtained. Analysis of the deduced amino acid (aa) sequence revealed two complete open reading frames (ORFs) (ORF1 and ORF3) and a truncated ORF (ORF4'). ORF1 has no homology to sequences in the databases, whereas ORF3 and ORF4' have significant homology to several bacterial DnaA (replication initiator) and DnaE (DNA polymerase III) proteins respectively. RT-PCR data showed that orf1-gyrB are co-transcribed, while dnaA-dnaE are co-transcribed but in the opposite direction. These data indicated that the gene organization of the Td gyrB region is unique compared with that of other bacteria. Eighteen putative DnaA boxes with several AT-rich regions were identified in the dnaA-dnaE intergenic region, and three putative DnaA boxes were identified in the gyrB-dnaA intergenic region. Spontaneous coumermycin A(1)-resistant Td mutants were isolated and characterized. The mutants have a >20-fold higher resistance to coumermycin A(1) than wild-type Td. A single point mutation in gyrB that changed GyrB Lys(136) to Glu or Thr appears to be responsible for the coumermycin A(1) resistance.
Program in Infectious Diseases, Department of Epidemiology, School of Public Health, University of North Carolina, Chapel Hill, NC 27599-7400, USA. srgreene@email.unc.edu
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D000595:Amino Acid Sequence; D049933:Aminocoumarins; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D016384:Consensus Sequence; D003374:Coumarins / Q000494:pharmacology; D027081:DNA Gyrase; D004258:DNA Polymerase III / Q000235:genetics; D004250:DNA Topoisomerases, Type II / Q000235:genetics*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics; D004352:Drug Resistance, Microbial; D008969:Molecular Sequence Data; D009154:Mutation; D016366:Open Reading Frames / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D014208:Treponema / Q000187:drug effects / Q000235:genetics*
D049933:Aminocoumarins; D001426:Bacterial Proteins; D003374:Coumarins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria; C074904:DNA polymerase III, alpha subunit; D004258:DNA Polymerase III; D027081:DNA Gyrase; D004250:DNA Topoisomerases, Type II; C004628:coumermycin