An unusual mechanism for resistance to the antibiotic coumermycin A1.
Bacterial DNA gyrases are type II topoisomerases made up of two A subunits and two B subunits. Coumarins are carbohydrate-containing antibiotics that inhibit topoisomerases II by competing with ATP for binding to the enzymes. High resistance to coumarins is produced in bacterial species by mutations in gyrB, the gene encoding subunit B. We have found an unusual mechanism of resistance to coumarins in Escherichia coli. This mechanism is exhibited by cells containing the wild-type gyrB, or its 5' half, in high copy number. Since homologous mutant gyrB (coumermycin resistant) truncated genes did not confer drug resistance at all under the same conditions, we propose that this mechanism of resistance is due to drug sequestration by the overproduced wild-type GyrB polypeptides. A corollary of this is that the amino half of GyrB is required and sufficient to fashion the ATP-binding domain of DNA gyrase, a conclusion that was further supported by mapping three independent coumarin-resistant mutations at Arg-136 of GyrB. Just upstream of this residue there is a glycine-rich sequence highly conserved in all topoisomerases II, which seems to be a good candidate for the actual ATP-binding site.
del Castillo
I
I
Unidad de Genética Molecular, Hospital Ramón y Cajal, Madrid, Spain.
Vizán
J L
JL
Rodríguez-Sáinz
M C
MC
Moreno
F
F
D000595:Amino Acid Sequence; D049933:Aminocoumarins; D003001:Cloning, Molecular; D003062:Codon; D003374:Coumarins / Q000494:pharmacology; D004252:DNA Mutational Analysis; D004250:DNA Topoisomerases, Type II / Q000502:physiology*; D004269:DNA, Bacterial / Q000235:genetics; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000187:drug effects*; D015870:Gene Expression; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D010957:Plasmids; D015183:Restriction Mapping
D049933:Aminocoumarins; D003062:Codon; D003374:Coumarins; D004269:DNA, Bacterial; D004250:DNA Topoisomerases, Type II; C004628:coumermycin