Binding of 3'-anthraniloyl-2'-deoxy-ATP to calmodulin-activated adenylate cyclase from Bordetella pertussis and Bacillus anthracis. - Institut Pasteur Accéder directement au contenu
Article Dans Une Revue Journal of Biological Chemistry Année : 1990

Binding of 3'-anthraniloyl-2'-deoxy-ATP to calmodulin-activated adenylate cyclase from Bordetella pertussis and Bacillus anthracis.

Résumé

3'-Anthraniloyl-2'-deoxyadenosine 5'-triphosphate (Ant-dATP), a fluorescent analogue of ATP, was tested as a probe for the nucleotide-binding site of calmodulin (CaM)-activated adenylate cyclases from Bordetella pertussis (BPCYA47) and Bacillus anthracis (BACYA62). Ant-dATP competitively inhibited both bacterial enzymes expressed in Escherichia coli (ki approximately 10 microM). Binding of the analogue to adenylate cyclase was monitored by equilibrium dialysis and by an increase in its fluorescence emission at 420 nm upon excitation at 330 nm. Whereas the fluorescence of Ant-dATP was little influenced by divalent cations, CaM, or adenylate cyclase alone, the Ca2+.CaM.cyclase complex increased up to 4 times the quantum yield of Ant-dATP. Binding of the analogue to the catalytic site of BPCYA47 and BACYA62 was specific as shown by its displacement with ATP or 3'-dATP. Our results substantiate the role of CaM in favoring substrate binding to CaM-activated enzymes.
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Dates et versions

pasteur-00167102 , version 1 (14-08-2007)

Identifiants

  • HAL Id : pasteur-00167102 , version 1
  • PUBMED : 2172237

Citer

Robert S. Sarfati, Vinod K. Kansal, Hélène Munier, Philippe Glaser, Anne-Marie Gilles, et al.. Binding of 3'-anthraniloyl-2'-deoxy-ATP to calmodulin-activated adenylate cyclase from Bordetella pertussis and Bacillus anthracis.. Journal of Biological Chemistry, 1990, 265 (31), pp.18902-6. ⟨pasteur-00167102⟩

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