INOSITOL 1,4,5-TRISPHOSPHATE AFFINITY-CHROMATOGRAPHY

Inositol 1,4,5-trisphosphate (IP3) affinity columns were made by coupling IP3 analogs to a supporting matrix, Sepharose 4B. IP3 5-phosphatase activity, IP3 3-kinase activity and IP3 binding activity from rat brain were adsorbed on the IP3 columns, and were eluted by increasing KCl concentration. This purification procedure increased the specific activities of these parameters 5-200-fold. Thus Sepharose 4B immobilized IP3 analogs can specifically interact with IP3-binding proteins, demonstrating that IP3 affinity columns are a good method for purifying such proteins. Furthermore, our results suggest that IP3 analogs can be linked to other molecules to make useful derivatives without loss of their biological activities. © 1990.