Synaptotagmin is an inositol polyphosphate binding protein: isolation and characterization as an Ins 1,3,4,5-P4 binding protein.
We isolated a binding protein for inositol 1,3,4,5-tetrakisphosphate (InsP4) from detergent-solubilized mouse cerebellar membrane fractions by sequential column chromatographies. Partial amino acid sequencing of the purified sample revealed that the protein is essentially identical to rat synaptotagmin II, an integral membrane protein of synaptic vesicles. Immunoprecipitation experiment of [3H]InsP4 binding activity of the purified protein using polyclonal antibody against the C2A domain of rat synaptotagmin II also revealed that mouse synaptotagmin II is the InsP4 binding protein (IP4BP). Scatchard analysis of InsP4 binding to the IP4BP/synaptotagmin indicates a single binding site with a Kd of 30 nM. The present finding that InsP4 binds strongly to synaptotagmin II suggests an important role for inositol polyphosphates in the regulation of neurotransmitter release.
Niinobe
M
M
Institute for Protein Research, Osaka University, Japan.
Yamaguchi
Y
Y
Fukuda
M
M
Mikoshiba
K
K
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002531:Cerebellum / Q000378:metabolism*; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D007295:Inositol Phosphates / Q000378:metabolism / Q000494:pharmacology; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D051379:Mice; D008815:Mice, Inbred Strains; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D017386:Sequence Homology, Amino Acid; D050857:Synaptotagmins
D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D007295:Inositol Phosphates; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; C081655:inositol-1,3,4,5-tetrakisphosphate; D050857:Synaptotagmins