Characterization of NAADP⁺ binding in sea urchin eggs

Nicotinic acid adenine dinucleotide phosphate (NAADP⁺) is a pyridine nucleotide which has been shown to release Ca²⁺ from intracellular membranes in echinoderms, Ascidiae, mammals, and plants. NAADP releases Ca²⁺ via a mechanism independent of ryanodine and inositol 1,4,5-trisphosphate (IP₃) 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. (C) 2000 Academic Press.