Inositol 1,3,4,5-tetrakisphosphate and inositol hexakisphosphate receptor proteins: Isolation and characterization from rat brain

High-affinity, membrane-associated inositol 1,3,4,5-tetrakisphosphate (IP₄) and inositol hexakisphosphate (IP₆) binding proteins were solubilized and isolated utilizing a heparin-agarose resin followed by an IP₄ affinity resin. The IP₆ receptor comprises a protein complex of 115-, 105-, and 50-kDa subunits, all of which comigrate under native conditions. The K_d of the receptor for IP₆ is 12 nM, whereas inositol 1,3,4,5,6-pentakisphosphate (IP₅), IP₄, and inositol 1,4,5-trisphosphate (IP₃) are 50%, 30%, and 15%, respectively, as potent. Two protein complexes copurify with the IP₄ receptor fraction. A 182/123-kDa complex elutes first from the affinity column followed by a 174/84-kDa protein complex, which elutes at higher salt. Both complexes show high affinity for IP₄ (K_d = 3-4 nM). IP₅, IP₆, and IP₃ display approximately 25%, 10%, and 0.1%, respectively, the affinity of IP₄. Ligand binding to IP₆ and IP₄ receptors is inhibited 50% by heparin at 0.1 μg/ml. IP₄ receptor proteins are stoichiometrically phospborylated by cyclic AMP-dependent protein kinase and protein kinase C, whereas negligible phosphorylation is observed for the IP₆ receptor. (.