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Extraction of major acidic Ca 2+ dependent phosphoproteins from synaptic membranes
Author(s) -
PerroneBizzozero N. I.,
Weiner D.,
Hauser G.,
Benowitz L. I.
Publication year - 1988
Publication title -
journal of neuroscience research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.72
H-Index - 160
eISSN - 1097-4547
pISSN - 0360-4012
DOI - 10.1002/jnr.490200308
Subject(s) - membrane , ionic strength , chemistry , extraction (chemistry) , chromatography , phosphatidylinositol , guanidine , gel electrophoresis , amphiphile , biochemistry , biophysics , phosphorylation , biology , organic chemistry , aqueous solution , copolymer , polymer
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 Ca 2+ 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.