Structure-activity relationships for the interactions of 2′- and 3′-(O)-(N-methyl)anthraniloyl-substituted purine and pyrimidine nucleotides with mammalian adenylyl cyclases

Cibele Pinto, Gerald H. Lushington, Mark Richter, Andreas Gille, Jens Geduhn, Burkhard König, Tung Chung Mou, Stephen R. Sprang, Roland Seifert

Research output: Contribution to journalArticlepeer-review

Abstract

Membranous adenylyl cyclases (ACs) play a key role in signal transduction and are promising drug targets. In previous studies we showed that 2′,3′-(O)-(N-methylanthraniloyl) (MANT)-substituted nucleotides are potent AC inhibitors. The aim of this study was to provide systematic structure-activity relationships for 21 (M)ANT-substituted nucleotides at the purified catalytic AC subunit heterodimer VC1:IIC2, the VC1:VC1 homodimer and recombinant ACs 1, 2 and 5. (M)ANT-nucleotides inhibited fully activated VC1:IIC2 in the order of affinity for bases hypoxanthine > uracil > cytosine > adenine ∼ guanine xanthine. Omission of a hydroxyl group at the 2′ or 3′-position reduced inhibitor potency as did introduction of a γ-thiophosphate group or omission of the γ-phosphate group. Substitution of the MANT-group by an ANT-group had little effect on affinity. Although all nucleotides bound to VC1:IIC2 similarly according to the tripartite pharmacophore model with a site for the base, the ribose, and the phosphate chain, nucleotides exhibited subtle differences in their binding modes as revealed by fluorescence spectroscopy and molecular modelling. MANT-nucleotides also differentially interacted with the VC1:VC1 homodimer as assessed by fluorescence spectroscopy and modelling. Similar structure-activity relationships as for VC1:IIC2 were obtained for recombinant ACs 1, 2 and 5, with AC2 being the least sensitive AC isoform in terms of inhibition. Overall, ACs possess a broad base-specificity with no preference for the "cognate" base adenine as verified by enzyme inhibition, fluorescence spectroscopy and molecular modelling. These properties of ACs are indicative for ligand-specific conformational landscapes that extend to the VC1:VC1 homodimer and should facilitate development of non-nucleotide inhibitors.

Original languageEnglish
Pages (from-to)358-370
Number of pages13
JournalBiochemical Pharmacology
Volume82
Issue number4
DOIs
StatePublished - Aug 15 2011

Keywords

  • Adenylyl Cyclase
  • Conformational landscape
  • Fluorescence spectroscopy
  • MANT-nucleotides
  • Molecular modelling

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