Interaction of nucleolar protein B23 with peptides related to nuclear localization signals.
Nucleolar protein B23 is a putative ribosome assembly factor with a high affinity for peptides containing nuclear localization signals (NLSs). The interactions of various NLS-containing peptides with two B23 isoforms (B23.1 and B23.2) were examined using equilibrium dialysis and Scatchard analyses. The KD for protein B23 binding to a peptide containing the SV40 T-antigen NLS sequence was approximately 1 microM with a stoichiometry of 1:1 (peptide:protein). No significant differences were seen between the two B23 isoforms in their affinities for any of the peptides tested. Binding by a reverse sequence SV40 T-NLS peptide showed a nonlinear Scatchard plot: this peptide was unable displace the correct sequence peptide, suggesting that the reverse sequence peptide binds to a different site on the protein. A peptide containing the sequence required for nucleolar localization of the HIV-1 Rev protein had an affinity for B23 approximately 10-fold greater than that of the SV40 T-NLS. However, with a sequence sufficient only for Rev location in the nucleoplasm, the affinity for B23 was diminished to a level between that of the longer Rev sequence and the SV40 T-NLS. In competition binding assays, the Rev NLS peptide was able to displace the SV40 T NLS, indicating that both peptides bind to the same site on protein B23. There was no detectable binding to protein B23 by a peptide containing the bipartite NLS of nucleoplasmin. Phosphorylation of protein B23 by casein kinase II enhanced its affinity for the SV40 T- and Rev-derived peptides approximately 2-fold.(ABSTRACT TRUNCATED AT 250 WORDS)
Szebeni
A
A
Department of Biochemistry, University of Mississippi Medical Center, Jackson 39216-4505, USA.
Herrera
J E
JE
Olson
M O
MO
D000595:Amino Acid Sequence; D000952:Antigens, Polyomavirus Transforming / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D047390:Casein Kinase II; D003956:Dialysis; D005033:Ethylmaleimide; D015694:Gene Products, rev / Q000737:chemistry / Q000378:metabolism; D015497:HIV-1 / Q000737:chemistry; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D000090243:Nucleophosmin; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D054321:rev Gene Products, Human Immunodeficiency Virus
D000952:Antigens, Polyomavirus Transforming; D015694:Gene Products, rev; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D054321:rev Gene Products, Human Immunodeficiency Virus; D000090243:Nucleophosmin; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D005033:Ethylmaleimide