Specific binding of cyclic ADP-ribose to calcium-storing microsomes from sea urchin eggs.

摘要: Cyclic ADP-ribose (cADPR) is a metabolite of NAD+ which as active inositol trisphosphate (IP3) in mobilizing intracellular Ca2+ sea urchin eggs. The enzyme responsible for synthesizing cADPR found not only eggs but also various mammalian tissue extracts, suggesting that it may be general messenger mobilization cells. In this study I address questions whether an receptor exists and, if so, different from the IP3 receptor. A procedure employing nitrogen decompression was used to homogenize eggs, and Ca2(+)-storing microsomes were separated mitochondria other organelles by Percoll density centrifugation. Radioactive with high specific activity produced incubating [32P]NAD+ product purified pressure liquid chromatography. membrane bound isolated dog brain extracts sucrose gradient Partial purification achieved DEAE ion-exchange chromatography after solubilization 3-[(cholamidopropyl)dimethylammonio]-1-propanesulfonate. Specific binding 32P-labeled saturable site on detected filtration assay. Scatchard analysis indicated affinity about 17 nM capacity 25 fmol/mg protein. affected either (the precursor) or hydrolysis product) at 0.5 microM eliminated 0.3 nonlabeled cADPR. appeared since effective competitor concentration 3 microM. Similarly, heparin inhibits most IP3-induced calcium release did affect binding. showed prominent pH optimum 6.7. Calcium 40 decreased 50%. These dependencies are those reported provide further support receptors different.