Serotonin-binding proteins in the bovine cerebral cortex: interaction with serotonin and catecholamines.
The soluble serotonin-binding proteins (SBP) present in bovine frontal cortex are very similar to those reported in rat brain. Binding of [3H]serotonin to SBP, present in ammonium sulphate-precipitated proteins from bovine cortex, requires Fe2+ but not Fe3+. In the presence of an optimal concentration of Fe2+ (0.1 mM), bovine SBP behave as a single class of non-cooperative sites for [3H]serotonin binding (Bmax = 120 +/- 12 pmol/mg protein, KD = 0.12 +/- 0.04 microM, n = 3). Binding of [3H]serotonin is decreased by nucleotides and by reagents which modify sulfhydryl groups and reduce disulfide bonds and by metal ion chelators. Serotonin analogs possessing an hydroxyl group on the indole ring and catecholamine analogs possessing an intact catechol moiety are effective competitors (Ki from 0.1 to 0.3 microM). In both cases, the aliphatic amino group does not contribute to the binding, but the affinity is strongly decreased if aromatic hydroxyl groups are methoxylated. Catecholamine-SBP interactions can also be demonstrated directly by binding experiments. Binding of [3H]dopamine is greatly enhanced by Fe2+, Cu2+ and Mn2+, but not by Fe3+. The Fe(2+)-dependent binding component of [3H]dopamine is saturable (Bmax = 279 +/- 64 pmol/mg protein, KD = 0.19 +/- 0.02 microM, n = 3), and possesses the same physicochemical properties as SBP: it elutes immediately after the void volume on a Sephacryl S100 HR (1.6 x 140 cm) gel filtration column (reflecting aggregation) and it migrates with an apparent molecular weight of 57-58 kDa on native polyacrylamide gel electrophoresis. Whereas the serotonin-storing role of SBP in serotonergic neurons has already been well documented, the present data advocate that these proteins may also possess catecholamine-storing properties.
Jimenez Del Rio
M
M
Department of Protein Chemistry, Free University Brussels, St. Genesius-Rode, Belgium.
Pinxteren
J
J
De Potter
W
W
Ebinger
G
G
Vauquelin
G
G
D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002395:Catecholamines / Q000378:metabolism*; D002417:Cattle; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D007202:Indicators and Reagents; D007501:Iron / Q000378:metabolism; D008970:Molecular Weight; D011485:Protein Binding; D012701:Serotonin / Q000378:metabolism*
D002352:Carrier Proteins; D002395:Catecholamines; D002614:Chelating Agents; D007202:Indicators and Reagents; C025366:serotonin-binding protein; D012701:Serotonin; D007501:Iron