Characterization of a high-affinity Ins-P4(inositol 1,3,4,5-tetrakisphosphate) receptor from brain by an anti-peptide antiserum

Abstract

From a high‐affinity Ins‐P₄ (inositol 1,3,4,5‐P₄) receptor purified from pig cerebellum, digested with the protease Lys C peptide sequences were obtained. Synthetic peptide‐3 (19 amino acid residues) was used to generate an antiserum. Reaction of the affinity‐purified antibodies with the purified pig receptor protein in ELISA or Western blot was completely inhibited by peptide‐3. In cerebellar membranes, the antibodies clearly recognized the 42 kDa Ins‐P₄ receptor protein and two additional proteins (25 kDa, 37 kDa) which still have to be identified. The anti‐peptide antibodies could selectively immunoprecipitate the Ins‐P₄ receptor protein. The antiserum was used (i) to demonstrate that in brain from different species (human, pig, beef, rat, mouse and sheep) a similar 42 kDa Ins‐P₄ receptor protein is contained, and (ii) to obtain indications for the existence of a related soluble form of the 42 kDa Ins‐P₄ receptor besides the membrane‐associated receptor.