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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

1,4-Dithiothreitol-induced alteration in histamine H1-agonist binding in guinea-pig cerebellum and cerebral cortex.

The effect of the disulphide bond reducing agent 1,4-dithiothreitol (DTT) on the binding characteristics of the H1-selective ligand [3H]mepyramine has been examined in homogenates of guinea-pig cerebral cortex and cerebellum. DTT was found to be without effect on antagonist binding. This was evident from studies using different concentrations of [3H]mepyramine (0.5-10.0 nM), which showed no change in either the equilibrium dissociation constant (KD) or specific binding site capacity (Bmax). Furthermore, the IC50 values and slope parameters determined from the inhibition of the binding of 1 nM [3H]mepyramine by non-radioactive mepyramine were similarly insensitive to DTT in both cerebral cortex and cerebellum. In contrast, DTT shifted the inhibition curve for histamine to lower agonist concentrations and reduced the Hill coefficient in these two tissues. Analysis of these inhibition curves as double hyperbolae revealed two binding sites in the presence of DTT and only one low affinity site in the absence of DTT. Similar changes in the location (IC50) and slope (Hill coefficient) parameters were obtained with the H1-selective agonist 2-thiazolylethylamine in cerebellum and with 2-methylhistamine in both brain regions. The results of this study show that DTT affects agonist but not antagonist binding in guinea-pig cerebellum and cerebral cortex, and suggest that DTT stabilises a proportion of the agonist binding sites in a high affinity state.[1]


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