Identification of a triplex DNA-binding protein from human cells.
Intramolecular or intermolecular triple helices could be recognized by specific proteins that stabilize triplex structures and might play a role in gene regulation. In order to identify such proteins, we designed a 55 nucleotide-long DNA oligomer that could fold on itself to form an intramolecular triple helix of the Py Pu x Py motif. The stability of this triplex under physiological conditions was demonstrated by gel retardation and thermal denaturation experiments. We have identified a protein from HeLa cell nuclear extracts that binds to this synthetic oligonucleotide. The protein has an apparent molecular mass of 55 kDa. Electrophoretic mobility shift assays revealed that the protein did not have any affinity for the single-stranded and double-stranded oligonucleotides corresponding respectively to the third strand and the Watson-Crick duplex of the triple helix. This protein also binds to an intramolecular Py Pu x Pu triplex but with a lower affinity than to a Py Pu x Py triple helix.
Guieysse
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AL
Laboratoire de Biophysique INSERM U.201 - CNRS URA 481, Muséum National d'Histoire Naturelle, Paris, France.
Praseuth
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D
Hélène
C
C
D001482:Base Composition; D001483:Base Sequence; D001973:Bromodeoxyuridine / Q000737:chemistry / Q000378:metabolism; D004247:DNA / Q000138:chemical synthesis / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000378:metabolism; D011089:Polydeoxyribonucleotides / Q000378:metabolism; D013696:Temperature; D014466:Ultraviolet Rays
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D011089:Polydeoxyribonucleotides; C051802:triplex DNA; C045717:poly d(C-T); D004247:DNA; D001973:Bromodeoxyuridine