Glyceraldehyde-3-phosphate dehydrogenase and Nm23-H1/nucleoside diphosphate kinase A. Two old enzymes combine for the novel Nm23 protein phosphotransferase function.
We have recently discovered an alternative function of the putative metastasis suppressor protein Nm23, which is identical to nucleoside diphosphate kinase, as a protein phosphotransferase in vitro. While purified native Nm23 protein did not phosphorylate other proteins, we could purify a Nm23-associated protein that activates the protein phosphotransferase function; it was identified as a glyceraldehyde-3-phosphate dehydrogenase (GAPDH) isoenzyme. Co-expression and purification of (His)6-tagged GAPDH in combination with either Nm23-H1 or Nm23-H2 in baculovirus-infected Sf9 cells showed that only Nm23-H1, but not Nm23-H2, forms a stable complex with GAPDH. Protein phosphotransferase activity was confirmed for the recombinant GAPDH.Nm23-H1 complex but not for either of the enzymes alone, nor was this activity observed after simple mixing of the purified proteins in vitro. The molecular mass of the highly purified recombinant GAPDH.Nm23-H1 complex suggests that a dimer of GAPDH interacts with a dimer of Nm23-H1. In contrast to the complex with GAPDH, co-expression of Nm23-H1 with antioxidant protein (MER-5) or creatine kinase did not activate the protein phosphotransferase function, indicating that this activation may specifically require GAPDH as a binding partner.
Engel
M
M
Department of Human Genetics, University of Saarland, D-66421 Homburg, Germany.
Seifert
M
M
Theisinger
B
B
Seyfert
U
U
Welter
C
C
D003402:Creatine Kinase / Q000378:metabolism; D019281:Dimerization; D004789:Enzyme Activation / Q000502:physiology; D004912:Erythrocytes / Q000201:enzymology; D015870:Gene Expression / Q000235:genetics; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D006801:Humans; D020559:Monomeric GTP-Binding Proteins; D054778:NM23 Nucleoside Diphosphate Kinases; D009363:Neoplasm Proteins / Q000378:metabolism; D009701:Nucleoside-Diphosphate Kinase / Q000502:physiology*; D010544:Peroxidases; D061105:Peroxiredoxin III; D054464:Peroxiredoxins; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000502:physiology*
D054778:NM23 Nucleoside Diphosphate Kinases; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D010544:Peroxidases; C516115:PRDX3 protein, human; D061105:Peroxiredoxin III; D054464:Peroxiredoxins; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010770:Phosphotransferases; D003402:Creatine Kinase; C517938:NME1 protein, human; D009701:Nucleoside-Diphosphate Kinase; D020559:Monomeric GTP-Binding Proteins