title: A nucleoside diphosphate kinase A (nm23-H1) serine 120-->glycine substitution in advanced stage neuroblastoma affects enzyme stability and alters protein-protein interaction.
abstract: A high level of nucleoside diphosphate kinase A (NDPK A/nm23-H1) in neuroblastoma is associated with advanced stage disease. We have also found a serine 120-->glycine substitution in NDPK A and/or amplification of the nm23-H1 gene in advanced stage neuroblastomas. Serine 120, a highly conserved residue, is located in proximity to histidine 118 which forms a phosphorylated intermediate essential for NDPK activity. The effect of Ser120-->Gly substitution on the biochemical properties of NDPK A was investigated. Phosphate-transferase activity was lower in the recombinant mutant NDPK A and in the immunoprecipitated complex consisting of NDPK A and NDPK B prepared from a neuroblastoma tumor containing the mutation, relative to the wild-type. There was a significant decrease in the enzyme stability toward urea- or temperature-induced denaturation for the recombinant mutant NDPK A and in an immunoprecipitate from a tumor containing the mutation. Recombinant NDPK A containing the Ser120-->Gly mutation exhibited reduced hexameric and increased dimeric oligomerization relative to the wild-type. Moreover a 28 kDa cellular protein was detected, that co-precipitated with the mutant but not wild-type NDPK A. The altered properties of the mutant protein may have relevance to a role for NDPK A in neuroblastoma progression.
authors.affiliation: Department of Pediatrics, University of Michigan, Ann Arbor 48109-0510, USA.
authors.forename: C L
authors.initials: CL
authors.lastname: Chang
authors.forename: J R
authors.initials: JR
authors.lastname: Strahler
authors.forename: D H
authors.initials: DH
authors.lastname: Thoraval
authors.forename: M G
authors.initials: MG
authors.lastname: Qian
authors.forename: R
authors.initials: R
authors.lastname: Hinderer
authors.forename: S M
authors.initials: SM
authors.lastname: Hanash
mesh_terms: D001483:Base Sequence; D003432:Cross-Linking Reagents; D017931:DNA Primers; D004795:Enzyme Stability; D005976:Glutaral; D005998:Glycine; D006358:Hot Temperature; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D020559:Monomeric GTP-Binding Proteins; D016297:Mutagenesis, Site-Directed; D054778:NM23 Nucleoside Diphosphate Kinases; D009367:Neoplasm Staging; D009447:Neuroblastoma / Q000201:enzymology* / Q000235:genetics* / Q000473:pathology; D009701:Nucleoside-Diphosphate Kinase / Q000737:chemistry / Q000378:metabolism; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012694:Serine; D013816:Thermodynamics; D014157:Transcription Factors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D003432:Cross-Linking Reagents; D017931:DNA Primers; D007527:Isoenzymes; D054778:NM23 Nucleoside Diphosphate Kinases; D011994:Recombinant Proteins; D014157:Transcription Factors; D012694:Serine; C517938:NME1 protein, human; D009701:Nucleoside-Diphosphate Kinase; D020559:Monomeric GTP-Binding Proteins; D005976:Glutaral; D005998:Glycine