Characterization of NAADP(+) binding in sea urchin eggs

Abstract

Nicotinic acid adenine dinucleotide phosphate (NAADP(+)) is a pyridine nucleotide which has been shown to release Ca(2+) from intracellular membranes in echinoderms, Ascidiae, mammals, and plants. NAADP releases Ca(2+) via a mechanism independent of ryanodine and inositol 1,4,5-trisphosphate (IP(3)) receptors and the NAADP(+) receptor is likely to be located on a separate organelle. We have investigated the binding characteristics of NAADP(+) to its receptor in sea urchin egg homogenates. NAADP(+) binds to a saturable membrane-bound site with high affinity (K(d) = 193 +/- 35. 7 pM). NAADP(+) associates to its receptor with a t(1/2) of approximately 7 min while dissociation does not occur during the time course of the experiment. Furthermore, NAD(+), NAAD(+), ADP, or ATP cannot displace NAADP(+) binding. The structurally related molecules NADP(+) and NADPH displayed a markedly lower affinity for the binding site with K(d)'s 500- and 25,000-fold higher than NAADP(+), respectively. This discrepancy between oxidized and reduced forms of NADP(+) might suggest that NAADP(+) signaling is itself regulated by the redox state of the cell.