title: Extraction of major acidic Ca2+ dependent phosphoproteins from synaptic membranes.
abstract: The association of several phosphoproteins with the synaptosomal plasma membrane (SPM) was investigated by phosphorylating SPM fractions from neonatal rat brain in the presence of Ca2+ and then exposing these to a variety of agents. Extraction of the major acidic phosphoproteins, GAP-43, pp40, and pp80, was assessed by two-dimensional gel electrophoresis and fluorography. All three proteins were best extracted from the membrane by high pH and by guanidine hydrochloride. GAP-43 was not extracted in the presence of either low- or high-ionic-strength buffers, reducing agents, or chelating agents; pp80 and pp40, however, showed a significant extraction even under low-ionic-strength conditions. Partition experiments with Triton X-114 revealed an amphiphilic behavior for GAP-43 and a strong affinity for hydrophobic environments for pp80 and pp40. None of the phosphoproteins was released from the membrane by the use of a phosphatidylinositol-specific phospholipase C. The extraction properties of GAP-43, pp80, and pp40 are similar to those of known extrinsic membrane proteins and therefore suggest that these phosphoproteins are peripheral rather than integral to the membrane compartment.
authors.affiliation: Mailman Research Center, McLean Hospital, Belmont, Massachusetts 02178.
authors.forename: N I
authors.initials: NI
authors.lastname: Perrone-Bizzozero
authors.forename: D
authors.initials: D
authors.lastname: Weiner
authors.forename: G
authors.initials: G
authors.lastname: Hauser
authors.forename: L I
authors.initials: LI
authors.lastname: Benowitz
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D001923:Brain Chemistry; D019922:GAP-43 Protein; D008565:Membrane Proteins / Q000302:isolation & purification*; D009419:Nerve Tissue Proteins / Q000302:isolation & purification*; D010750:Phosphoproteins / Q000302:isolation & purification*; D051381:Rats; D013574:Synaptosomes / Q000032:analysis*
chemical_list: D019922:GAP-43 Protein; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins