Nucleic acid-binding properties of the Xenopus oocyte Y box protein mRNP3+4.
Y box proteins contain the conserved cold shock domain (CSD) and several basic/aromatic (B/A) islands that are rich in arginine and aromatic residues. The binding of purified Xenopus oocyte 6S Y box protein, mRNP3+4, to Y box RNA, single-stranded (ss) DNA, and double-stranded (ds) DNA was studied by gel mobility shift and nitrocellulose filter binding assays. mRNP3+4 specifically bound Y box ssDNA or RNA, while binding of dsDNA was not detected. Y box ssDNA and RNA did not efficiently cross-compete for mRNP3+4 binding, and no evidence for ternary complex formation was detected. However, Y box ssDNA binding was competed by high concentrations of Y box RNA or nonspecific RNA competitors, indicating that the ssDNA-binding site has a lower affinity for RNA. mRNP3+4 demonstrated similar affinity for either Y box RNA or ssDNA. However, at elevated ionic strength RNA binding was markedly greater than ssDNA binding, indicating that RNA binding involves nonionic interactions that are not utilized for ssDNA binding. Recombinant polypeptides containing B/A islands bound Y box RNA exclusively, but inclusion of the CSD led to preferential ssDNA binding. The results demonstrate that the B/A islands are exclusively RNA-binding, while the CSD exhibits preferential binding of ssDNA. The inability of Y box RNA and ssDNA to efficiently cross-compete for mRNP3+4 binding suggests that isoforms exhibit preferential ssDNA or RNA binding.
Murray
M T
MT
Center for Molecular Biology, Wayne State University, Detroit, Michigan 48202.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004277:DNA, Single-Stranded / Q000378:metabolism*; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism*; D011485:Protein Binding; D012313:RNA / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D014981:Xenopus; D029867:Xenopus Proteins
D004277:DNA, Single-Stranded; D016601:RNA-Binding Proteins; D014157:Transcription Factors; D029867:Xenopus Proteins; C075593:YBX2 protein, Xenopus; D012313:RNA