Evidence for multiple forms of melatonin receptor-G-protein complexes by solubilization and gel electrophoresis.
The daily production of melatonin from the pineal gland influences circadian and seasonal behaviour and physiology. To further understand how melatonin may function, it is important to characterize the receptor and signal transduction systems. Using the detergent digitonin, we were able to solubilize the receptor from the ovine pars tuberalis (PT) membrane. The receptor was isolated as a complex associated with its heterotrimeric G-protein. In the solubilized state, pre-bound 125I-2-iodomelatonin was stable at 4 degrees C, but was displaceable by GTP gamma S. The receptor-G-protein complex could be separated by molecular mass using native polyacrylamide gel electrophoresis. We demonstrate that the receptor-complex has a molecular mass of 525 kDa and differs from solubilized receptor-complexes isolated from either the lizard brain, chicken brain or the ovine hippocampus. Furthermore the receptor complex isolated from the hippocampus had the lowest molecular mass of these tissues (365 kDa) and was found not to be sensitive to GTP gamma S. This may indicate the existence of a distinct non-G-protein coupled form of the receptor.
Barrett
P
P
Rowett Research Institute, Bucksburn, Aberdeen, Scotland, UK.
MacLean
A
A
Morgan
P J
PJ
D000818:Animals; D001923:Brain Chemistry; D002645:Chickens; D004072:Digitonin; D004591:Electrophoresis, Polyacrylamide Gel; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006624:Hippocampus / Q000737:chemistry; D007457:Iodine Radioisotopes; D008116:Lizards; D046911:Macromolecular Substances; D008550:Melatonin / Q000378:metabolism; D008970:Molecular Weight; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism*; D044094:Receptors, Melatonin; D012756:Sheep; D015398:Signal Transduction; D012995:Solubility
D007457:Iodine Radioisotopes; D046911:Macromolecular Substances; D011956:Receptors, Cell Surface; D044094:Receptors, Melatonin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019204:GTP-Binding Proteins; D008550:Melatonin; D004072:Digitonin