Purification of the heat-stable inhibitor protein of the Ca2+-activated cyclic nucleotide phosphodiesterase by affinity chromatography.
The recently discovered heat-stable inhibitor protein of the Ca2+-activated cyclic nucleotide phosphodiesterase (Sharma, R. K., Wirch, E. & Warg, J. H. (1978) J. Biol. Chem., in press) has been purified 238 214-fold from bovine brain extract using an affinity column of the modulator protein--Sepharose 4B conjugate. The purified sample appears to be homogeneous as judged by sodium dodecyl sulphate (SDS) gel electrophoresis. The protein band has a mobility corresponding to that of a polypeptide of molecular weight 68 000. Since the heat-stable inhibitor protein has a molecular weight of 70 000 under nondenaturing conditions, it suggests that it is a monomeric protein. The protein has no inhibitory activity toward the cAMP-dependent protein kinase or protein phosphatase. The purified sample has been tested for various enzyme activities which include ATPase, GTPase, cAMP phosphodiesterase, cGMP phosphodiesterase, 5'-nucleotidase, and protein kinase. None of these activities are exhibited by the purified sample.
D015087:2',3'-Cyclic-Nucleotide Phosphodiesterases / Q000037:antagonists & inhibitors / Q000378:metabolism; D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D001923:Brain Chemistry; D002118:Calcium / Q000378:metabolism*; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods; D006358:Hot Temperature; D009419:Nerve Tissue Proteins / Q000302:isolation & purification; D011494:Protein Kinases / Q000378:metabolism
D002352:Carrier Proteins; D009419:Nerve Tissue Proteins; D011494:Protein Kinases; D015087:2',3'-Cyclic-Nucleotide Phosphodiesterases; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D002118:Calcium