Purified inositol 1,4,5-trisphosphate receptor mediates calcium flux in reconstituted lipid vesicles.
Inositol 1,4,5-trisphosphate (Ins(1,4,5)P3), a second messenger molecule involved in actions of neurotransmitters, hormones and growth factors, releases calcium from vesicular non-mitochondrial intracellular stores. An Ins(1,4,5)P3 binding protein, purified from brain membranes, has been shown to be phosphorylated by cyclic-AMP-dependent protein kinase and localized by immunohistochemical techniques to intracellular particles associated with the endoplasmic reticulum. Although the specificity of the Ins(1,4,5)P3 binding protein for inositol phosphates and the high affinity of the protein for Ins(1,4,5)P3 indicate that it is a physiological Ins(1,4,5)P3 receptor mediating calcium release, direct evidence for this has been difficult to obtain. Also, it is unclear whether a single protein mediates both the recognition of Ins(1,4,5)P3 and calcium transport or whether these two functions involve two or more distinct proteins. In the present study we report reconstitution of the purified Ins(1,4,5)P3 binding protein into lipid vesicles. We show that Ins(1,4,5)P3 and other inositol phosphates stimulate calcium flux in the reconstituted vesicles with potencies and specificities that match the calcium releasing actions of Ins(1,4,5)P3. These results indicate that the purified Ins(1,4,5)P3 binding protein is a physiological receptor responsible for calcium release.
Ferris
C D
CD
Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Huganir
R L
RL
Supattapone
S
S
Snyder
S H
SH
D000818:Animals; D001667:Binding, Competitive; D001921:Brain / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels; D002793:Cholic Acids; D002846:Chromatography, Affinity; D003902:Detergents; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism / Q000502:physiology*; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007700:Kinetics; D008081:Liposomes; D008566:Membranes / Q000378:metabolism; D008970:Molecular Weight; D010713:Phosphatidylcholines; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D018160:Receptors, Cytoplasmic and Nuclear; D015290:Second Messenger Systems
D015220:Calcium Channels; D002793:Cholic Acids; D003902:Detergents; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008081:Liposomes; D010713:Phosphatidylcholines; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D000001:Calcimycin; D015544:Inositol 1,4,5-Trisphosphate; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate; D002118:Calcium