Purification of the growth-associated protein GAP-43 by reversed phase chromatography: amino acid sequence analysis and cDNA identification.
GAP-43 is a neuronal phosphoprotein. Increased synthesis and axonal transport of GAP-43 has been associated with axon growth, and altered phosphorylation of GAP-43 has been associated with changes in synaptic efficacy. Here we report a rapid and effective procedure employing reverse-phase HPLC for the purification of GAP-43 from rat brain. To characterize the protein purified by this procedure, we generated proteolytic fragments and determined their amino acid sequences. These directly determined sequences, corresponding to 56% of the GAP-43 amino acids, confirm recently reported sequences deduced from the nucleotide sequences of cDNAs. Using oligonucleotide probes constructed according to these amino acid sequences, we identified GAP-43 cDNAs in a library prepared from neonatal rat superior cervical ganglion cells. One of these cDNAs was 1.1 kB in size; it hybridized specifically with a 1.5 kB RNA from brain, but not from liver, and contained the entire coding sequence for GAP-43. This cDNA differed from recently reported cDNAs in its 3' untranslated region.
Changelian
P S
PS
Department of Anatomy and Neurobiology, Washington University School of Medicine, St. Louis, MO 63110.
Meiri
K
K
Soppet
D
D
Valenza
H
H
Loewy
A
A
Willard
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000032:analysis*; D019922:GAP-43 Protein; D008562:Membrane Glycoproteins / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D009414:Nerve Growth Factors / Q000235:genetics / Q000302:isolation & purification*; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000302:isolation & purification*; D009693:Nucleic Acid Hybridization; D051381:Rats
D019922:GAP-43 Protein; D008562:Membrane Glycoproteins; D009414:Nerve Growth Factors; D009419:Nerve Tissue Proteins; D004247:DNA