[Identification of the cardiac beta-adrenergic receptor protein: solubilization and purification by affinity chromatography].

ABSTRACT A protein that binds catecholamines with a specificity parallel to that of their in vivo effects on cardiac contractility (isoproterenol > epinephrine or norepinephrine > dopamine > dihydroxyphenylalanine) was solubilized from a microsomal fraction of canine ventricular myocardium. The binding protein was purified 500 to 800-fold by solubilization and subsequent affinity chromatography with conjugates of norepinephrine linked to agarose beads. Purified beta-adrenergic binding protein exists in two forms, corresponding to molecular weights of 40,000 and 160,000. The purified material has a single association constant, 2.3 x 10(5) liters/mol (as compared to two association constants, 10(7) and 10(6) liters/mol, for the binding protein in particulate form) but retains the identical binding specificity for beta-adrenergic drugs and antagonists.

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    Biochemical pharmacology 09/2013; DOI:10.1016/j.bcp.2013.09.011 · 4.65 Impact Factor
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    ABSTRACT: Detergent extracts of isolated rat liver plasma membranes were analysed in two-dimensional immunoelectrophoresis against antiserum to plasma membranes. Enzyme staining of the immuno-precipitates revealed the presence of about ten antigens with nucleoside di- and triphosphatase activity. Most of these were earlier shown also to be NADH-neotetrazolium reductase active. In addition, two of these antigens exhibited l-leucyl-β-naphthylamidase activity.As judged from autoradiography these plasma membrane antigens earlier characterized as multienzyme complexes bound [14C]epinephrine, and the same antigens were labelled regardless of whether membranes or membrane extracts were incubated with the radioactive hormone. The specificity of this binding was established in displacement experiments with unlabelled hormones or their analogues.Another hormone-binding antigen, also identified in the plasma membrane extract did not exhibit any known enzyme activity while three antigens with different enzyme activities had no epinephrine-binding capacity.[14C]Epinephrine-labelled plasma membrane extracts were chromatographed on Sepharose 4B and the fractions obtained were analysed in two-dimensional immunoelectrophoresis combined with autoradiography. Nucleoside di- and triphosphatases of high molecular weights (500000) were associated with l-leucyl-β-naphthylamidase activity, while no such associations were detected in a lower molecular weight region (70000). Further immunological studies on the various fractionated antigens provided evidence that at least two of them occurred in both low and high molecular weight fractions. Hormone-binding membrane components in varying concentrations were found throughout the eluted extract.
    08/1975; 56(2):319-326. DOI:10.1111/j.1432-1033.1975.tb02236.x
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    ABSTRACT: The conformational structure of propranolol, a β-adrenergic blocking drug, has been investigated by pcilo calculations and 270-MHz proton nuclear magnetic resonance in D2O solution. The molecules coexist in at least two conformational states in solution with a low energy barrier. Both preferred conformations have extended structures which allow a three-point drug-receptor binding involving the aromatic moiety, the β-hydroxyl group, and -NH+2 groups of propranolol. The previously postulated “rigid” bicyclic structure does not exist to an appreciable extent in D2O solution.
    International Journal of Quantum Chemistry 03/2009; 16:153-170. DOI:10.1002/qua.560160711 · 1.17 Impact Factor

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