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Design of an Affinity Matrix for Purification of the Histamine H 1 Receptor from Guinea Pig Cerebellum
Author(s) -
Ruat M.,
Traiffort E.,
Garbarg M.,
Schwartz J. C.,
Demonchaux P.,
Ife R. J.,
Tertiuk W.,
Ganellin C. R.
Publication year - 1992
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1992.tb09317.x
Subject(s) - mepyramine , digitonin , sepharose , guinea pig , cerebellum , chemistry , histamine , radioligand assay , receptor , binding site , glycine , stereochemistry , chromatography , amino acid , biochemistry , antagonist , enzyme , biology , neuroscience , endocrinology
H 1 receptors from guinea pig cerebellum were solubilized using digitonin, and [ 125 I]iodobolpyramine was used as a probe. [ 125 I]Iodobolpyramine binding to this solubilized preparation occurred with a K D of 0.1 n M and a B max of 220 fmol/mg of protein and was inhibited by various H 1 ligands with the expected potencies. Using a gel filtration procedure, a very sensitive radioassay was set up for detecting H 1 activity in the solubilized preparation: 0.1 n M [ 125 I]iodobolpyramine specific binding represented >90% of total binding. Moreover, the synthesis is described of potent H 1 antagonists that are mepyramine derivatives with an amino alkyl acylamido alkyl spacer arm. One of them, UCL 1057 ( K i = 0.5 n M ), has been coupled to a Sepharose epoxy‐activated resin. The resulting affinity matrix adsorbed selectively [ 125 I]iodobolpyramine binding sites from the guinea pig cerebellum soluble preparation. In contrast, a Sepharose–glycine matrix was not able to adsorb these sites.