Differences in phosphorylation of human and chicken stathmin by MAP kinase.
Stathmin/Op18 is a highly conserved 19 kDa cytosolic phosphoprotein. Human and chicken stathmin share 93% identity with only 11 amino acid substitutions. One of the substituted amino acids is serine 25, which is a glycine in chicken stathmin. In human stathmin, serine 25 is the main phosphorylation site for MAP kinase. In this study, we have compared the phosphorylation of human and chicken stathmin. The proteins were expressed in Sf9 cells using the baculovirus expression system and purified for in vitro phosphorylation assays. Phosphorylation with MAP kinase showed that chicken stathmin was phosphorylated 10 times less than human stathmin. To identify the phosphorylation sites we used liquid chromatography/mass spectrometry (LC/MS/MS). The only amino acid found phosphorylated was serine 38, which corresponds to the minor phosphorylation site in human stathmin. Phosphorylation with p34(cdc2)- and cGMP-dependent protein kinases gave almost identical phosphorylation levels in the two stathmins.
Antonsson
B
B
Geneva Biomedical Research Institute, Glaxo Wellcome R&D S.A., 14 ch des Aulx, CH-1288 Plan-les-Ouates, Geneva, Switzerland. bruno.antonsson.ch_gva03@serono.com
Kassel
D B
DB
Ruchti
E
E
Grenningloh
G
G
D000595:Amino Acid Sequence; D000818:Animals; D006801:Humans; D013058:Mass Spectrometry; D008868:Microtubule Proteins; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010750:Phosphoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D050777:Stathmin
D008868:Microtubule Proteins; D010750:Phosphoproteins; D011994:Recombinant Proteins; C493995:STMN1 protein, human; D050777:Stathmin; D020928:Mitogen-Activated Protein Kinases