A mass spectrometric study on the in vivo posttranslational modification of GAP-43

J Biol Chem. 1994 Sep 9;269(36):22481-4.

Abstract

GAP-43 isolated from calf brain was analyzed by the electrospray mass spectrometry. The mass spectrum of the intact protein showed two species with a mass difference of 80 Da, suggesting that the isolated GAP-43 contains phosphorylated species. To establish the in vivo phosphorylation sites, the protein was digested with trypsin, and analyzed by the liquid chromatography/mass spectrometry technique, in which a capillary reversed-phase chromatography column was connected on line to an electrospray mass spectrometer. Two pairs of peptides with a mass difference of 80 Da were observed. From the tandem mass spectrometry, two novel phosphorylation sites (Thr-87 and Ser-152) were identified. The novel phosphorylation sites contain proline immediately after the phosphorylated serines. No phosphorylated peptide was detected corresponding to the protein kinase C or casein kinase II phosphorylation sites. A peptide corresponding to the acetylated N-terminal peptide was also identified. The mass of the peptide suggests that the 2 cysteinyl residues are not palmitoylated but form a disulfide bridge.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Brain / metabolism
  • Cattle
  • Chromatography, Affinity
  • Chromatography, High Pressure Liquid
  • Chromatography, Ion Exchange
  • GAP-43 Protein
  • Intracellular Signaling Peptides and Proteins*
  • Mass Spectrometry / methods
  • Membrane Glycoproteins / biosynthesis*
  • Membrane Glycoproteins / chemistry*
  • Membrane Glycoproteins / isolation & purification
  • Membrane Proteins*
  • Molecular Sequence Data
  • Myristoylated Alanine-Rich C Kinase Substrate
  • Nerve Tissue Proteins / biosynthesis*
  • Nerve Tissue Proteins / chemistry*
  • Nerve Tissue Proteins / isolation & purification
  • Peptide Fragments / chemistry
  • Peptide Fragments / isolation & purification
  • Phosphopeptides / chemistry
  • Phosphopeptides / isolation & purification
  • Phosphoproteins / chemistry*
  • Protein Processing, Post-Translational*
  • Proteins / chemistry
  • Trypsin

Substances

  • GAP-43 Protein
  • Intracellular Signaling Peptides and Proteins
  • Membrane Glycoproteins
  • Membrane Proteins
  • Nerve Tissue Proteins
  • Peptide Fragments
  • Phosphopeptides
  • Phosphoproteins
  • Proteins
  • Myristoylated Alanine-Rich C Kinase Substrate
  • Trypsin