Solubilization and hydrophobic immobilization assay of a cAMP binding protein from Dictyostelium discoideum plasma membranes |
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Authors: | B L Meyers W A Frazier |
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Institution: | Department of Biological Chemistry Division of Biology and Biomedical Sciences Washington University School of Medicine St. Louis, MO 63110 USA |
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Abstract: | A cAMP binding site present on isolated plasma membranes of aggregation-competent cells has been solubilized with the nonionic detergent Emulphogene BC-720. An assay has been developed based on the principle of hydrophobic chromatography, in which the detergent solubilized cAMP binding protein is immobilized on alkyl-agarose beads at low detergent concentration. This allows the necessary rapid separation of bound and free 3H]-cAMP by filtration of the beads. The kinetics and nucleotide specificity of the detergent solubilized cAMP binding protein are comparable to those of the cAMP chemotactic receptor on intact cells and plasma membranes. The alkyl-agarose bead assay may have general utility for the assay of detergent solubilized membrane receptors. |
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Keywords: | membrane phosphodiesterase mPDE bovine serum albumin BSA |
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