Affinity chromatography with the immobilized agonist N-0434 yields an active and highly purified preparation of the dopamine D-2 receptor from bovine striatum.
Partial purification of the dopamine D-2 receptor from bovine striatum, solubilized in the presence of 1% digitonin, was obtained by chromatography on wheat germ lectin agarose. The preparation was purified approximately 10-fold. The stability of the receptor preparation was considerably improved and non-specific protein absorption on the affinity gel used later was decreased. Further purification was achieved on a column containing a D-2-selective agonist, N-0434. Approximately 90% of the receptor activity was bound to the gel and 20-40% of the activity could be eluted by pH shock. The total purification factor after one affinity chromatography step was estimated to be at least 1500. An active preparation of at least 20% purity was obtained after a second cycle of affinity chromatography. This corresponds to an enrichment of more than 5000 times compared to the solubilized receptor preparation.
Bosker
F J
FJ
Department of Medical Biochemistry, Sylvius Laboratories, University of Leiden, The Netherlands.
Van Bussel
F J
FJ
Thielen
A P
AP
Soei
Y L
YL
Sieswerda
G T
GT
Dijk
J
J
Tepper
P G
PG
Horn
A S
AS
Möller
W
W
D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity; D003342:Corpus Striatum / Q000187:drug effects / Q000378:metabolism*; D015259:Dopamine Agents; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D037102:Lectins; D009419:Nerve Tissue Proteins / Q000378:metabolism; D010627:Phenethylamines; D011954:Receptors, Dopamine / Q000187:drug effects*; D013134:Spiperone / Q000378:metabolism
D015259:Dopamine Agents; D037102:Lectins; D009419:Nerve Tissue Proteins; D010627:Phenethylamines; D011954:Receptors, Dopamine; D013134:Spiperone; C041369:2-(N-phenethyl-N-propyl)amino-5-hydroxytetralin