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. 1991 Apr;10(4):885–892. doi: 10.1002/j.1460-2075.1991.tb08021.x

Identification of the regulatory phosphorylation sites in pp42/mitogen-activated protein kinase (MAP kinase).

D M Payne 1, A J Rossomando 1, P Martino 1, A K Erickson 1, J H Her 1, J Shabanowitz 1, D F Hunt 1, M J Weber 1, T W Sturgill 1
PMCID: PMC452730  PMID: 1849075

Abstract

Mitogen-activated protein kinase (MAP kinase) is a 42 kd serine/threonine protein kinase whose enzymatic activity requires phosphorylation of both tyrosyl and threonyl residues. As a step in elucidating the mechanism(s) for activation of this enzyme, we have determined the sites of regulatory phosphorylation. Following proteolytic digestion of 32P-labeled pp42/MAP kinase with trypsin, only a single phosphopeptide was detected by two-dimensional peptide mapping, and this peptide contained both phosphotyrosine and phosphothreonine. The amino acid sequence of the peptide, including the phosphorylation sites, was determined using a combination of Fourier transform mass spectrometry and collision-activated dissociation tandem mass spectrometry with electrospray ionization. The sequence for the pp42/MAP kinase tryptic phosphopeptide is similar (but not identical) to a sequence present in the ERK1- and KSS1-encoded kinases. The two phosphorylation sites are separated by only a single residue. The regulation of activity by dual phosphorylations at closely spaced threonyl and tyrosyl residues has a functional correlate in p34cdc2, and may be characteristic of a family of protein kinases regulating cell cycle transitions.

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Selected References

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