1. Academic Validation
  2. Analogues of the nicotinic acid adenine dinucleotide phosphate (NAADP) antagonist Ned-19 indicate two binding sites on the NAADP receptor

Analogues of the nicotinic acid adenine dinucleotide phosphate (NAADP) antagonist Ned-19 indicate two binding sites on the NAADP receptor

  • J Biol Chem. 2009 Dec 11;284(50):34930-4. doi: 10.1074/jbc.M109.016519.
Daniel Rosen 1 Alexander M Lewis Akiko Mizote Justyn M Thomas Parvinder K Aley Sridhar R Vasudevan Raman Parkesh Antony Galione Minoru Izumi A Ganesan Grant C Churchill
Affiliations

Affiliation

  • 1 Department of Pharmacology, University of Oxford, Mansfield Road, Oxford OX1 3QT, United Kingdom.
Abstract

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a CA(2+)-releasing messenger. Biological data suggest that its receptor has two binding sites: one high-affinity locking site and one low-affinity opening site. To directly address the presence and function of these putative binding sites, we synthesized and tested analogues of the NAADP antagonist Ned-19. Ned-19 itself inhibits both NAADP-mediated CA(2+) release and NAADP binding. A fluorometry bioassay was used to assess NAADP-mediated CA(2+) release, whereas a radioreceptor assay was used to assess binding to the NAADP receptor (only at the high-affinity site). In Ned-20, the fluorine is para rather than ortho as in Ned-19. Ned-20 does not inhibit NAADP-mediated CA(2+) release but inhibits NAADP binding. Conversely, Ned-19.4 (a methyl ester of Ned-19) inhibits NAADP-mediated CA(2+) release but cannot inhibit NAADP binding. Furthermore, Ned-20 prevents the self-desensitization response characteristic of NAADP in sea urchin eggs, confirming that this response is mediated by a high-affinity allosteric site to which NAADP binds in the radioreceptor assay. Collectively, these data provide the first direct evidence for two binding sites (one high- and one low-affinity) on the NAADP receptor.

Figures
Products