Isolation and characterization of vascular smooth muscle inositol 1,4,5-trisphosphate receptor

Abstract

myo-Inositol 1,4,5-trisphosphate (InsP₃) receptor of porcine aorta was purified to near homogeneity and its biochemical properties were compared with those of cerebellar InsP₃ receptor of the same animal species. The aortic InsP₃ receptor consisted of equal amounts of two polypeptides with slightly differing molecular masses of around 240 kDa and was found to possess a single population of InsP₃-binding site (K_d of 1.2 nM). The InsP₃ receptor purified from porcine cerebellum was also comprised of two polypeptides. However, the molecular mass was slightly but definitely larger, being 250 kDa, and the amounts of the two polypeptides were not equal. The aortic InsP₃ receptor cross-reacted with polyclonal antibody specific to type 1 InsP₃ receptor as did the cerebellar InsP₃ receptor. The aortic InsP₃ receptor bound to calmodulin–Sepharose in a Ca²⁺-dependent manner, while the cerebellar InsP₃ receptor did not. Reverse transcriptase-PCR analysis revealed two splicing variants of the type 1 InsP₃ receptor in porcine aortic smooth muscle distinct from those of the type 1 InsP₃ receptor of porcine cerebellum. The possible relevance of this difference to difference in calmodulin-binding property was discussed.