Purification of D2-dopamine receptors from bovine brain

1986 ◽  
Vol 14 (6) ◽  
pp. 1138-1139 ◽  
Author(s):  
SIMON WORRALL ◽  
RICHARD A. WILLIAMSON ◽  
PHILIP G. STRANGE
Keyword(s):  
Dopamine Receptors ◽  
Bovine Brain ◽  
D2 Dopamine Receptors

scholarly journals The glycosylation properties of D2 dopamine receptors from striatal and limbic areas of bovine brain

1988 ◽  
Vol 255 (3) ◽  
pp. 877-883 ◽  
Author(s):  
M N Leonard ◽  
R A Williamson ◽  
P G Strange
Keyword(s):  
Caudate Nucleus ◽  
Dopamine Receptors ◽  
Sodium Cholate ◽  
Bovine Brain ◽  
Affinity Column ◽  
Olfactory Tubercle ◽  
D2 Dopamine Receptors ◽  
Neuraminidase Treatment ◽  
Membrane Bound ◽  
Affinity Interaction

D2 dopamine receptors from bovine brain (caudate nucleus and olfactory tubercle) have been solubilized using sodium cholate/NaCl and their glycoprotein properties studied in terms of their interaction with wheat-germ agglutinin-agarose (WGA-agarose). Under optimal conditions about 65% of the applied D2 dopamine receptors bound to WGA-agarose and could be eluted with N-acetylglucosamine. The ability of receptors to adsorb to the affinity column was shown to be dependent on the cholate and salt concentrations used. Digestion of the membrane bound D2 dopamine receptors with neuraminidase prior to solubilisation reduced the ability of the receptors to bind to WGA-agarose (50% of applied receptors bound) whereas digestion with N-acetylglucosaminidase did not significantly affect binding to WGA-agarose. Digestion with the two enzymes together resulted in a larger decrease in binding to WGA-agarose than was seen with the two enzymes alone (40% of applied receptors bound). Stepwise elution of bound receptors from the WGA-agarose columns using 2.5 mM- and 100-mM-N-acetylglucosamine showed that about 40% of the bound receptors interacted with WGA-agarose in a low-affinity manner, the remainder showing a high-affinity interaction. Neuraminidase treatment reduced the low-affinity population suggesting that the interaction of oligosaccharides bearing sialic acid with WGA-agarose is of lower affinity and that higher-affinity binding is via N-acetylglucosamine. These data are discussed in terms of the heterogeneity of carbohydrate moieties on the D2 dopamine receptors within a brain region. In all the tests applied here, however, receptors from caudate nucleus and olfactory tubercle behaved identically so their glycosylation patterns must be very similar.

scholarly journals Antisera specific for D2 dopamine receptors

1993 ◽  
Vol 289 (3) ◽  
pp. 789-794 ◽  
Author(s):  
P L Chazot ◽  
A J Doherty ◽  
P G Strange
Keyword(s):  
Dopamine Receptors ◽  
Cho Cells ◽  
Receptor Gene ◽  
Bovine Brain ◽  
Extracellular Loop ◽  
Intracellular Loop ◽  
D2 Dopamine Receptors ◽  
Western Blots ◽  
Recombinant Cho Cells ◽  
Third Intracellular Loop

Antisera have been raised against two peptides from the sequence of D2 dopamine receptors: peptide 1 from the predicted second extracellular loop and peptide 2 from the predicted third intracellular loop. The antisera recognize specifically a 95 kDa band in Western blots of several bovine brain regions, which corresponds to the denatured D2 dopamine receptor, whereas in recombinant CHO cells expressing D2 dopamine receptors a 80 kDa band is seen. The antisera immunoprecipitate 10-20% of the D2 dopamine receptors from soluble preparations of bovine brain. The antisera recognize D2 dopamine receptors in immunofluorescence analyses of recombinant CHO cells bearing the receptor gene. The antisera directed against the third intracellular loop, but not those against the second extracellular loop, will interfere with the coupling of D2 dopamine receptors and G-proteins in bovine brain preparations.

scholarly journals Heterogeneity of D2 dopamine receptors in different brain regions

1987 ◽  
Vol 248 (2) ◽  
pp. 595-602 ◽  
Author(s):  
M N Leonard ◽  
C A Macey ◽  
P G Strange
Keyword(s):  
Caudate Nucleus ◽  
Dopamine Receptors ◽  
Serotonin Receptors ◽  
Brain Regions ◽  
Bovine Brain ◽  
Olfactory Tubercle ◽  
Agonist Binding ◽  
D2 Dopamine Receptors ◽  
Substituted Benzamides ◽  
Nucleotide Regulation

The binding of [3H]spiperone has been examined in membranes derived from different regions of bovine brain. In caudate nucleus, nucleus accumbens, olfactory tubercle and putamen binding is to D2 dopamine and 5HT2 serotonin receptors, whereas in cingulate cortex only serotonin 5HT2 receptor binding can be detected. D2 dopamine receptors were examined in detail in caudate nucleus, olfactory tubercle and putamen using [3H]spiperone binding in the presence of 0.3 microM-mianserin (to block 5HT2 serotonin receptors). No evidence for heterogeneity among D2 dopamine receptors either between brain regions or within a brain region was found from the displacements of [3H]spiperone binding by a range of antagonists, including dibenzazepines and substituted benzamides. Regulation of agonist binding by guanine nucleotides did, however, differ between regions. In caudate nucleus a population of agonist binding sites appeared resistant to guanine nucleotide regulation, whereas this was not the case in olfactory tubercle and putamen.

Partial purification of D2 dopamine receptors from bovine brain

1987 ◽  
Vol 15 (6) ◽  
pp. 1155-1156 ◽  
Author(s):  
RICHARD A. WILLIAMSON ◽  
SIMON WORRALL ◽  
PHILIP G. STRANGE
Keyword(s):  
Dopamine Receptors ◽  
Bovine Brain ◽  
Partial Purification ◽  
D2 Dopamine Receptors
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