title: The histidine-221 to tyrosine substitution in v-mos abolishes its biological function and its protein kinase activity.
abstract: The viral mos gene encodes a cytoplasmic transforming protein termed p37mos. Evidence gathered from a number of experimental approaches is consistent with p37mos having a serine/threonine protein kinase activity. To gain further understanding of the p37mos-associated biochemical activity, we constructed a mutation in the v-mos gene by oligonucleotide-directed mutagenesis yielding a histidine to tyrosine substitution at residue 221 in p37mos. Based upon nucleotide sequences, the histidine residue at the corresponding position is conserved in all the serine/threonine protein kinases from yeast to man, and is absent in protein-tyrosine kinases. The mutant p37mos (Tyr-221) was expressed in yeast and assayed for kinase activity. The mutant protein was inactive as judged by a loss of autophosphorylation activity in vitro, thus providing further support for the conclusion that p37mos is a protein kinase. When the mutant v-mos gene was introduced into a retroviral vector, pDD102, and assayed for focus-forming ability on NIH/3T3 cells, it was found to be inactive at both 37 and 30 degrees. In contrast, the wild-type v-mos had transforming activity at both temperatures. These results extend our earlier findings on the correlation between transforming ability and protein kinase activity. A histidine to tyrosine substitution at the corresponding position of the v-mos protein and the yeast CDC28 gene product causes a similar effect on the kinase activity. Therefore, this residue and/or the sequence near the N-terminal side of the conserved predicted phosphate transfer domain, near the middle of the complete catalytic domain, might be specifically involved in the catalytic activity of serine/threonine protein kinases in general.
authors.affiliation: Department of Molecular Pathology, University of Texas System Cancer Center, M.D. Anderson Hospital and Tumor Institute, Houston 77030.
authors.forename: B
authors.initials: B
authors.lastname: Singh
authors.forename: C
authors.initials: C
authors.lastname: Wittenberg
authors.forename: M
authors.initials: M
authors.lastname: Hannink
authors.forename: S I
authors.initials: SI
authors.lastname: Reed
authors.forename: D J
authors.initials: DJ
authors.lastname: Donoghue
authors.forename: R B
authors.initials: RB
authors.lastname: Arlinghaus
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002472:Cell Transformation, Viral; D005347:Fibroblasts; D005656:Fungal Proteins / Q000235:genetics; D006639:Histidine / Q000502:physiology; D051379:Mice; D017232:Oncogene Proteins v-mos; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics* / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012191:Retroviridae Proteins / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid; D014443:Tyrosine
chemical_list: D005656:Fungal Proteins; D017232:Oncogene Proteins v-mos; D011993:Recombinant Fusion Proteins; D012191:Retroviridae Proteins; D014443:Tyrosine; D006639:Histidine; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases