Solubilization and Characterization of Opioid Binding Sites from Frog (Rana esculenta) Brain |
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Authors: | J Simon M Szücs S Benyhe A Borsodi P Zeman M Wollemann |
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Institution: | Institute of Biochemistry, Biological Research Center of the Hungarian Academy of Sciences, Szeged, Hungary |
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Abstract: | Active opioid receptors were solubilized from frog (Rana esculenta) brain membrane fractions by the use of 1% digitonin. It was found by kinetic as well as by equilibrium measurements that both the membrane and the solubilized fractions contain two binding sites. For the membrane preparations, KD values were 0.9 and 3.6 nM, and Bmax values were 293 and 734 fmol/mg protein. For the solubilized preparations, KD values were 0.4 and 2.6 nM, an Bmax values were 35 and 266 fmol/mg protein. The stereospecificity of the binding did not change during solubilization. Both the membrane-bound and the solubilized receptors showed weak binding of enkephalin and mu-specific drugs, suggesting that they are predominantly of the kappa-type. The membrane-bound and the soluble receptors showed the same distribution of subtypes, i.e., 70% kappa, 13% mu, and 17% delta for the membrane-bound and 71% kappa, 17% mu, and 12% delta for the soluble receptors. |
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Keywords: | Frog brain Opiate receptors Solubilization [3H]Naloxone binding Existence of subtypes |
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