A 47-kDa human nuclear protein recognized by antikinetochore autoimmune sera is homologous with the protein encoded by RCC1, a gene implicated in onset of chromosome condensation.
Several autoimmune sera from patients with Raynaud phenomenon decorated mammalian kinetochores and bound to a 47-kDa protein on immunoblots of nuclear lysates. Antibody affinity-purified from immunoblots of the 47-kDa band recognized kinetochores, but due to crossreaction with an 18-kDa protein, localization remains elusive. We used one of these sera to purify the antigen from HeLa cells synchronized in mitosis as a noncovalent complex with a 25-kDa protein. The antigen was released from DNA by intercalation with 25 mM chloroquine. Ion-exchange chromatography yielded the pure complex with an apparent molecular size of 68 kDa, which was separated into its components by gel filtration in 6 M guanidinium chloride. Upon two-dimensional gel electrophoresis the 47-kDa protein gave two main spots of pI 6.6 and 6.7, respectively. Posttranslational modification is indicated by additional antigenic spots, by lack of a free alpha-amino group, and by chromatographic behavior of peptides on reversed-phase chromatography. The amino acid sequence for 205 residues of the 47-kDa protein has been established. This sequence is highly homologous with the translated reading frame of RCC1, a gene reportedly involved in regulating onset of mammalian chromosome condensation.
Bischoff
F R
FR
Institute of Cell and Tumor Biology, Deutsches Krebsforschungszentrum, Heidelberg, Federal Republic of Germany.
Maier
G
G
Tilz
G
G
Ponstingl
H
H
D000595:Amino Acid Sequence; D001323:Autoantibodies / Q000276:immunology*; D001324:Autoantigens / Q000032:analysis*; D001327:Autoimmune Diseases / Q000097:blood / Q000276:immunology*; D002877:Chromosomes, Human / Q000502:physiology*; D005260:Female; D005455:Fluorescent Antibody Technique; D005796:Genes; D006367:HeLa Cells / Q000276:immunology; D006801:Humans; D008875:Middle Aged; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D011928:Raynaud Disease / Q000097:blood / Q000276:immunology*; D012689:Sequence Homology, Nucleic Acid
D001323:Autoantibodies; D001324:Autoantigens; D009687:Nuclear Proteins