Purification of cyclic 3',5'-nucleotide phosphodiesterase inhibitory protein by affinity chromatography on activator protein coupled to sepharose
- 10 January 1978
- journal article
- research article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 17 (1) , 120-126
- https://doi.org/10.1021/bi00594a017
Abstract
The Ca2+-dependent, reversible, interaction of cyclic[c]AMP phosphodiesterase with its activator was used to purify the [bovine brain] enzyme by affinity chromatography. Activator-dependent cAMP phosphodiesterase is only a minor component of the proteins specifically adsorbed in the presence of Ca2+ by the Ca2+-dependent activator protein coupled to Sepharose and subsequently released by [ethylenebis(oxyethylenenitrilo)]tetraacetic acid. The major protein component can be partially resolved from the enzyme by gel filtration on Sephadex G-200. This protein was purified to apparent homogeneity and shown to be composed of 2 polypeptide chains with molecular weights of 61,000 and 15,000, respectively. This protein is, by itself, devoid of phosphodiesterase activity and inhibits the activation of cAMP phosphodiesterase by its activator without affecting the basal activity. Thus, activation of cAMP phosphodiesterase by the Ca2+-dependent activator protein may be controlled by interactions with yet a 3rd component of the enzyme complex.Keywords
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