ALTERED STRUCTURE OF THE DNA DUPLEX RECOGNIZED BY YEAST TRANSCRIPTIONFACTOR REB1P

The Saccharomyces cerevisiae REB1 gene encodes a sequence-specific DNA binding protein that has been implicated in chromatin structure, tran scription regulation and transcription termination. Previous work has shown that the DNA sequence recognized by Reb1p contains an adenosine residue that is unusually reactive toward chemical modification by dim ethylsulfate and that methylation of this nucleoside increases the bin ding affinity of the Reb1p protein for its target. Prompted by these r esults, we determined the solution structure of the 13mer Reb1p DNA du plex recognition site d(GTCCGGGTAATGC). d(GCATTACCCGGAC) using 2D NMR, distance geometry and iterative 2D NOESY back-calculation structure r efinement. The distance geometry-refined molecule demonstrated an unus ual structure in the TAAT region of the sequence that was manifested i n cross-strand base stacking, as indicated by unusually strong NOE int eractions between H2 protons on three adjacent adenosine bases. This s tructure was compared to two published NMR studies of DNA duplexes con taining the related sequence TAAC. The Reb1p DNA structure does not sh ow the conformational mobility or the 'transient kink' at TpA steps ch aracteristic of the related TAAT-containing sequences.