PMID:1656457

Citation	del Castillo, I, Vizán, JL, Rodríguez-Sáinz, MC and Moreno, F (1991) An unusual mechanism for resistance to the antibiotic coumermycin A1.Proc. Natl. Acad. Sci. U.S.A. 88:8860-4
Abstract	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.
Links	PubMed
Keywords	Amino Acid Sequence; Aminocoumarins; Cloning, Molecular; Codon; Coumarins; DNA Mutational Analysis; DNA Topoisomerases, Type II; DNA, Bacterial; Drug Resistance, Microbial; Escherichia coli; Gene Expression; Genes, Bacterial; Molecular Sequence Data; Plasmids; Restriction Mapping
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Phenotype Annotations

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Phenotype of	Taxon Information	Genotype Information (if known)	Condition Information	OMP ID	OMP Term Name	ECO ID	ECO Term Name	Notes	Status
a mutation or genetic difference within a strain	Taxon: Escherichia coli Strain: K-12 Substrain: RYC1000 NCBI Taxon ID: 83333	Genotype of Reference Strain: gyrB+ Genotype of Experimental Strain : Multi-copy gyrB in RYC1010	Reference Condition: without coumermycin Experimental Condition: With Coumermycin					The plasmid pCID549 was used to increase gene dosage of gyrB causing increased resistance toward coumermycin A1.
a mutation or genetic difference within a strain	Taxon: E-coli Strain: K12 Substrain: RYC1010 NCBI Taxon ID: 83333	Genotype of Reference Strain: gyrB+ Genotype of Experimental Strain : gyrB(del)	Reference Condition:					- Unusual mechanism of Ecoli gene gyrB resistance to the antibody coumermycin. - Single mutations at Arg-136 resulted in high level resistance to coumermycin A1, which concludes that this position in the gyrB gene is essential for coumermycin binding. - Refer to fig 3 in the article to see the gyrB polypeptide and the positions where mutation took place.
a mutation or genetic difference within a strain	Taxon: E-coli Strain: K12 Substrain: RYC1010 NCBI Taxon ID: 83333	Genotype of Reference Strain: gyrB+ Genotype of Experimental Strain : gyrB(truncated)	Reference Condition: without coumermycin Experimental Condition: with coumermycin					Plasmid pCID511.25 was constructed by eliminating 3' end of gyrB (removal of PvuII fragment, or elimination of the last 168 nucleotides). The gene only conferred resistance to coumermycin at 30C but not at 42C.
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