The binding of cyclic [3H] AMP by bovine adrenal extracts was studied in different buffer systems; 50 mM-Tris/HCl, pH 7.5, buffer containing 4mM-EDTA (disodium salt) offered advantages such as low non-specific interference and high binding affinity. 2-Mercaptoethanol decreased, but EDTA and theophylline increased, the specificity of cyclic [3H]AMP binding as expressed by the displacement of cyclic [3H]AMP by a 1000-fold excess of unlabelled cyclic GMP. The study indicated heterogeneity of the binding material. Mg2+-dependent binding may be eliminated in Tris/EDTA buffer. The heterogeneity and the specificity of intracellular cyclic AMP binders and their dependence in environmental factors should be considered in the evaluation of cyclic AMP binding and second-messenger systems.
Effect of buffer composition on binding of adenosine 3′:5′-cyclic monophosphate by bovine adrenal extracts
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J Linkola, F Fyhrquist, T H Weber, L Puutula; Effect of buffer composition on binding of adenosine 3′:5′-cyclic monophosphate by bovine adrenal extracts. Biochem J 15 September 1977; 166 (3): 553–557. doi: https://doi.org/10.1042/bj1660553
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