A 4.2 kDa synthetic peptide as a potential probe to evaluate the antibacterial activity of coumarin drugs.
The coumarin antibiotics are potent inhibitors of DNA replication whose target is the enzyme DNA gyrase, an ATP-dependent bacterial type II topoisomerase. The coumarin drugs inhibit gyrase action by competitive binding to the ATP-binding site of DNA gyrase B protein. The production of new biologically active products has stimulated additional studies on coumarin-gyrase interactions. In this regard, a 4.2 kDa peptide mimic of DNA gyrase B protein from Escherichia coli has been designed and synthesized. The peptide sequence includes the natural fragment 131-146 (coumarin resistance-determining region) and a segment containing the gyrase-DNA interaction region (positions 753-770). The peptide mimic binds to novobiocin (Ka = 1.4+/-0.3 x 10(5) M(-1)), plasmid (Ka = 1.6+/-0.5 x 10(6) M(-1)) and ATP (Ka = 1.9+/-50.4 x 10(3) M(-1)), results previously found with the intact B protein. On the other hand, the binding to novobiocin was reduced when a mutation of Arg-136 to Leu-136 was introduced, a change previously found in the DNA gyrase B protein from several coumarin-resistant clinical isolates of Escherichia coli In contrast, the binding to plasmid and to ATP was not altered. These results suggest that synthetic peptides designed in a similar way to that described here could be used as mimics of DNA gyrase in studies which seek a better understanding of the ATP, as well as coumarin, binding to the gyrase and also the mechanism of action of this class of antibacterial drugs.
Departamento de Bioquímica e Tecnologia Química, Instituto de Química, Universidade Estadual Paulista, Araraquara, São Paulo, Brazil.
Andreza C
AC
Scatigno
Saulo S
SS
Garrido
Reinaldo
R
Marchetto
D000255:Adenosine Triphosphate / Q000737:chemistry; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D001667:Binding, Competitive; D003374:Coumarins / Q000737:chemistry / Q000494:pharmacology*; D004247:DNA / Q000737:chemistry; D027081:DNA Gyrase / Q000737:chemistry*; D004353:Drug Evaluation, Preclinical; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology; D015335:Molecular Probes; D009675:Novobiocin / Q000737:chemistry; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D059005:Topoisomerase II Inhibitors
C498233:4.2 kDa peptide; D000900:Anti-Bacterial Agents; D003374:Coumarins; D004791:Enzyme Inhibitors; D015335:Molecular Probes; D010455:Peptides; D059005:Topoisomerase II Inhibitors; D009675:Novobiocin; D000255:Adenosine Triphosphate; D004247:DNA; D027081:DNA Gyrase