Sequence-specific interaction of the HMG box proteins TCF-1 and SRY occurs within the minor groove of a Watson-Crick double helix.
The high mobility group I (HMG) box is proposed to mediate DNA binding in a novel group of transcription-regulating proteins. Two of these, the proteins encoded by the T cell-specific TCF-1 and the mammalian sex-determining gene SRY, carry a single HMG box with specificity for the heptamer motif A/T A/T C A A A G. We have now analysed the mode of interaction of the HMG boxes of TCF-1 and SRY with this motif. Methylation interference footprinting revealed that both HMG boxes contacted adenines on both strands in the minor groove, whereas no major groove guanine contacts were discerned. Diethylpyrocarbonate (DEPC) carbethoxylation interference footprinting of TCF-1 indicated the absence of major groove contacts on positions 5, 6 and 7 of the motif. Carbethoxylation interference was observed, however, on positions 2, 3 and 4 and to a lesser extent on position 1 in the major groove. Combined T----C and A----I substitution, which changes the surface of the major groove but leaves the minor groove intact, did not interfere with sequence-specific binding by TCF-1 and SRY. These observations indicate that recognition of the heptamer motif by the HMG boxes of the distantly related TCF-1 and SRY proteins predominantly occurs through nucleotide contacts in the minor groove.
van de Wetering
M
M
Department of Immunology, University Hospital, Utrecht, The Netherlands.
Clevers
H
H
D000478:Alkylation; D000818:Animals; D001482:Base Composition; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D004047:Diethyl Pyrocarbonate; D004742:Enhancer Elements, Genetic; D004926:Escherichia coli / Q000235:genetics; D006609:High Mobility Group Proteins / Q000378:metabolism*; D051821:Lymphoid Enhancer-Binding Factor 1; D008745:Methylation; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides; D015345:Oligonucleotide Probes; D011948:Receptors, Antigen, T-Cell / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D051876:Sex-Determining Region Y Protein; D051822:T Cell Transcription Factor 1; D013601:T-Lymphocytes / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; D006609:High Mobility Group Proteins; D051821:Lymphoid Enhancer-Binding Factor 1; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D015345:Oligonucleotide Probes; D011948:Receptors, Antigen, T-Cell; D011994:Recombinant Proteins; D051876:Sex-Determining Region Y Protein; D051822:T Cell Transcription Factor 1; D014157:Transcription Factors; D004247:DNA; D004047:Diethyl Pyrocarbonate