Analysis of co-crystal structures to identify the stereochemical determinants of the orientation of TBP on the TATA box.

Possible stereochemical determinants of the orientation of TBP on the TATA box are discussed using the crystal coordinates of TBP-TATA complexes, which have been determined by other groups. The C-terminal half of the TBP beta-sheet interacts with the TATA site of the DNA, and the N-terminal half with the A-rich site, so that the two sites with distinct curvatures produce a unique fit. Although chemical contacts take place between one side of the beta-sheet and the DNA minor groove, the interaction seems to be facilitated indirectly by the characteristics of the other side of the beta-sheet and the DNA major groove. Thus, Ala71, Leu162 and Pro190 differentiate the curvature of the beta-sheet in the N- and C-halves. The methyl positions in the DNA major groove modulate the bendability of the two DNA sites by using differences in the rolling capacity of TA and AT compared with PyT, and in the shifting capacity of AT compared with TT. The deformations of the first steps (TA and PyT) in the two sites are the largest and thus are important for the overall bending of the DNA. The differences between the two DNA sites are greatest at the second steps (AT and TT) and so these are important for determining the orientation of TBP.     

The amino-terminal tails of the core histones and the translational position of the TATA box determine TBP/TFIIA association with nucleosomal DNA.

We establish that the TATA binding protein (TBP) in the presence of TFIIA recognizes the TATA box in nucleosomal DNA dependent on the dissociation of the amino-terminal tails of the core histones from the nucleosome and the position of the TATA box within the nucleosome. We examine TBP/TFIIA access to the TATA box with this sequence placed in four distinct rotational frames with reference to the histone surface and at three distinct translational positions at the edge, side and dyad axis of the nucleosome. Under our experimental conditions, we find that the preferential translational position at which TBP/TFIIA can bind the TATA box is within linker DNA at the edge of the nucleosome and that binding is facilitated if contacts made by the amino-terminal tails of the histones with nucleosomal DNA are eliminated. TBP/TFIIA binding to DNA at the edge of the nucleosome occurs with the TATA box in all four rotational positions. This is indicative of TBP/TFIIA association directing the dissociation of the TATA box from the surface of the histone octamer.     

Beyond the "recognition code": structures of two Cys2His2 zinc finger/TATA box complexes

BACKGROUND: Several methods have been developed for creating Cys2His2 zinc finger proteins that recognize novel DNA sequences, and these proteins may have important applications in biological research and gene therapy. In spite of this progress with design/selection methodology, fundamental questions remain about the principles that govern DNA recognition. One hypothesis suggests that recognition can be described by a simple set of rules--essentially a "recognition code"--but careful assessment of this proposal has been difficult because there have been few structural studies of selected zinc finger proteins. RESULTS: We report the high-resolution cocrystal structures of two zinc finger proteins that had been selected (as variants of Zif268) to recognize a eukaryotic TATA box sequence. The overall docking arrangement of the fingers within the major groove of the DNA is similar to that observed in the Zif268 complex. Nevertheless, comparison of Zif268 and the selected variants reveal significant differences in the pattern of side chain-base interactions. The new structures also reveal side chain-side chain interactions (both within and between fingers) that are important in stabilizing the protein-DNA interface and appear to play substantial roles in recognition. CONCLUSIONS: These new structures highlight the surprising complexity of zinc finger-DNA interactions. The diversity of interactions observed at the protein-DNA interface, which is especially striking for proteins that were all derived from Zif268, challenges fundamental concepts about zinc finger-DNA recognition and underscores the difficulty in developing any meaningful recognition code.