Selective determinants of inositol 1,4,5-trisphosphate and adenophostin A interactions with type 1 inositol 1,4,5-trisphosphate receptors.

ABSTRACT: Adenophostin A (AdA) is a potent agonist of inositol 1,4,5-trisphosphate receptors (IP(3) R). AdA shares with IP(3) the essential features of all IP(3) R agonists, namely structures equivalent to the 4,5-bisphosphate and 6-hydroxyl of IP(3) , but the basis of its increased affinity is unclear. Hitherto, the 2'-phosphate of AdA has been thought to provide a supra-optimal mimic of the 1-phosphate of IP(3) .We examined the structural determinants of AdA binding to type 1 IP(3) R (IP(3) R1). Chemical synthesis and mutational analysis of IP(3) R1 were combined with (3) H-IP(3) binding to full-length IP(3) R1 and its N-terminal fragments, and Ca(2+) release assays from recombinant IP(3) R1 expressed in DT40 cells.Adenophostin A is at least 12-fold more potent than IP(3) in functional assays, and the IP(3) -binding core (IBC, residues 224-604 of IP(3) R1) is sufficient for this high-affinity binding of AdA. Removal of the 2'-phosphate from AdA (to give 2'-dephospho-AdA) had significantly lesser effects on its affinity for the IBC than did removal of the 1-phosphate from IP(3) (to give inositol 4,5-bisphosphate). Mutation of the only residue (R568) that interacts directly with the 1-phosphate of IP(3) decreased similarly (by ~30-fold) the affinity for IP(3) and AdA, but mutating R504, which has been proposed to form a cation-? interaction with the adenine of AdA, more profoundly reduced the affinity of IP(3) R for AdA (353-fold) than for IP(3) (13-fold).The 2'-phosphate of AdA is not a major determinant of its high affinity. R504 in the receptor, most likely via a cation-? interaction, contributes specifically to AdA binding.

SUBMITTER: Rossi AM 

PROVIDER: S-EPMC2998688 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature