title: Identification of the major synaptojanin-binding proteins in brain.
abstract: Synaptojanin is a nerve-terminal enriched inositol 5-phosphatase thought to function in synaptic vesicle endocytosis, in part through interactions with the Src homology 3 domain of amphiphysin. We have used synaptojanin purified from Sf9 cells after baculovirus mediated expression in overlay assays to identify two major synaptojanin-binding proteins in rat brain. The first, at 125 kDa, is amphiphysin. The second, at 40 kDa, is the major synaptojanin-binding protein detected, is highly enriched in brain, is concentrated in a soluble synaptic fraction, and co-immunoprecipitates with synaptojanin. The 40-kDa protein does not bind to a synaptojanin construct lacking the proline-rich C terminus, suggesting that its interaction with synaptojanin is mediated through an Src homology 3 domain. The 40-kDa synaptojanin-binding protein was partially purified from rat brain cytosol through a three-step procedure involving ammonium sulfate precipitation, sucrose density gradient centrifugation, and DEAE ion-exchange chromatography. Peptide sequence analysis identified the 40-kDa protein as SH3P4, a member of a novel family of Src homology 3 domain-containing proteins. These data suggest an important role for SH3P4 in synaptic vesicle endocytosis.
authors.affiliation: Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, Quebec H3A 2B4, Canada.
authors.forename: E
authors.initials: E
authors.lastname: de Heuvel
authors.forename: A W
authors.initials: AW
authors.lastname: Bell
authors.forename: A R
authors.initials: AR
authors.lastname: Ramjaun
authors.forename: K
authors.initials: K
authors.lastname: Wong
authors.forename: W S
authors.initials: WS
authors.lastname: Sossin
authors.forename: P S
authors.initials: PS
authors.lastname: McPherson
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001923:Brain Chemistry; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification*; D002478:Cells, Cultured; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D051381:Rats; D018411:Spodoptera; D018909:src Homology Domains
chemical_list: D002352:Carrier Proteins; D009419:Nerve Tissue Proteins; C075637:amphiphysin; C097702:synaptojanin; D010744:Phosphoric Monoester Hydrolases