Cloning and primary structure of Staphylococcus aureus DNA topoisomerase IV: a primary target of fluoroquinolones

Abstract

A 4.6 kb Staphylococcus aureus DNA fragment containing DNA gyrase-like genes (grlA and grlB) was cloned and sequenced. The proteins GrlA and GrlB exhibit more than 30% identity with E. coli DNA topoisomerase IV subunits and with the gyrase subunits from S. aureus and Escherichia coli. The combined E. coli cell extracts of GrlA and GrlB overproducing strains catalysed ATP-dependent relaxation and decatenation specific to DNA topoisomerase IV. The temperature-sensitive phenotype of Salmonella typhimurium parC and parE mutants was complemented by the S. aureus grlA and grlB genes, when the two genes were co-expressed. These results show that GrlA and GrlB are the subunits of S. aureus DNA topoisomerase IV. The GyrA subunit of DNA gyrase has been previously defined as a primary target of quinolones based on genetic and biochemical experiments essentially carried out in E. coli. Single-point mutations occurring in the 'quinolone resistance-determining region' (QRDR) of GyrA were found in bacteria exhibiting quinolone resistance, the most common mutation being a substitution of Ser-83 on the E. coli GyrA sequence. We analysed eight S. aureus fluoroquinolone-resistant clinical isolates and observed that mutations in the QRDR of GyrA are not present in the low-quinolone-resistant isolates. In contrast, Ser-80 of GrlA, which corresponds to Ser-83 of E. coli GyrA, is substituted to Phe or Tyr in both high- and low-quinolone-resistant isolates. We propose that DNA topoisomerase IV is a primary target of fluoroquinolones in S. aureus.

Persistence of Mycoplasma synoviae in hens after two enrofloxacin treatments and detection of mutations in the parC gene

“…The detection of a mutation in the ParC QRDR suggests that the parC gene might be the primary target of enrofloxacin in M. synoviae as described for M. bovirhinis [16] or Staphylococcus aureus (grlA) [11]. Previous studies have shown Topoisomerase IV to be the primary target of ciprofloxacin (analogous to enrofloxacin) in M. hominis [3].…”

Canadian Guidelines for the Management of Acute Exacerbations of Chronic Bronchitis

“…In S pneumoniae, low-level fluoroquinolone resistance to ciprofloxacin and levofloxacin has been associated with mutations in the QRDR of parC. However, high-level resistance requires additional mutations in the QRDR of gyrA (255)(256)(257)(258)(259). Decreased accumulation: Acquired fluoroquinolone resistance due to active efflux in S pneumoniae has been shown to be due to increased levels of expression of efflux pumps mediated by the pmrA gene (260).…”

Identification of DNA gyrase Subunit a Mutations Associated with Ciprofloxacin Resistance in Staphylococcus aureus Isolated from Nasal Infection in Kurdistan-Iran

“…Amino acid replacement correlating with fluoroquinolone resistance in GrlA include Ser80->Phe or Tyr, Ala116->Glu or Pro, and Glu84->Lys (Kaatz & Seo, 1997;Agents an chemotherapy, 1992;Ferrero et al, 1994;Ferrero et al, 1995;Yamagishi et al, 1996). With respect to GyrA, Ser84->Leu or Ala, Ser85->Pro and Glu88->Lys mutations are associated with fluoroquinolone resistance (Kaatz & Seo, 1997;Goswitz et al, 1992;Kaatz & Seo, 1995) amino acid replacement in GyrB correlating with fluoroquinolone resistance include Asp437->Asn and Arg458->Glu (Ito et al, 1994;Kaatz & Seo, 1997).…”