Geometric analysis and comparison of protein-DNA interfaces: why is there no simple code for recognition?

Structural studies of protein-DNA complexes have shown that there are many distinct families of DNA-binding proteins, and have shown that there is no simple "code" describing side-chain/base interactions. However, systematic analysis and comparison of protein-DNA complexes has been complicated by the diversity of observed contacts, the sheer number of complexes currently available and the absence of any consistent method of comparison that retains detailed structural information about the protein-DNA interface. To address these problems, we have developed geometric methods for characterizing the local structural environment in which particular side-chain/base interactions are observed. In particular, we develop methods for analyzing and comparing spatial relationships at the protein-DNA interface. Our method involves attaching local coordinate systems to the DNA bases and to the C(alpha) atoms of the peptide backbone (these are relatively rigid structural units). We use these tools to consider how the position and orientation of the polypeptide backbone (with respect to the DNA) helps to determine what contacts are possible at any given position in a protein-DNA complex. Here, we focus on base contacts that are made in the major groove, and we use spatial relationships in analyzing: (i) the observed patterns of side-chain/base interactions; (ii) observed helix docking orientations; (iii) family/subfamily relationships among DNA-binding proteins; and (iv) broader questions about evolution, altered specificity mutants and the limits for the design of new DNA-binding proteins. Our analysis, which highlights differences in spatial relationships in different complexes and at different positions in a complex, helps explain why there is no simple, general code for protein-DNA recognition.

DNA recognition by Cys2His2 zinc finger proteins.

Cys2His2 zinc fingers are one of the most common DNA-binding motifs found in eukaryotic transcription factors. These proteins typically contain several fingers that make tandem contacts along the DNA. Each finger has a conserved beta beta alpha structure, and amino acids on the surface of the alpha-helix contact bases in the major groove. This simple, modular structure of zinc finger proteins, and the wide variety of DNA sequences they can recognize, make them an attractive framework for attempts to design novel DNA-binding proteins. Several studies have selected fingers with new specificities, and there clearly are recurring patterns in the observed side chain-base interactions. However, the structural details of recognition are intricate enough that there are no general rules (a "recognition code") that would allow the design of an optimal protein for any desired target site. Construction of multifinger proteins is also complicated by interactions between neighboring fingers and the effect of the intervening linker. This review analyzes DNA recognition by Cys2His2 zinc fingers and summarizes progress in generating proteins with novel specificities from fingers selected by phage display.

Zif268 protein-DNA complex refined at 1.6 A: a model system for understanding zinc finger-DNA interactions.

BACKGROUND: Zinc fingers of the Cys2 His2 class recognize a wide variety of different DNA sequences and are one of the most abundant DNA-binding motifs found in eukaryotes. The previously determined 2.1 A structure of a complex containing the three zinc fingers from Zif268 has served as a basis for many modeling and design studies, and Zif268 has proved to be a very useful model system for studying how TFIIIA-like zinc fingers recognize DNA. RESULTS: We have refined the structure of the Zif268 protein-DNA complex at 1.6 A resolution. Our structure confirms all the basic features of the previous model and allows us to focus on some critical details at the protein-DNA interface. In particular, our refined structure helps explain the roles of several acidic residues located in the recognition helices and shows that the zinc fingers make a number of water-mediated contacts with bases and phosphates. Modeling studies suggest that the distinctive DNA conformation observed in the Zif268-DNA complex is correlated with finger-finger interactions and the length of the linkers between adjacent fingers. Circular dichroism studies indicate that at least some of the features of this distinctive DNA conformation are induced upon complex formation. CONCLUSIONS: Our 1.6 A structure should provide an excellent framework for analyzing the effects of Zif268 mutations, for modeling related zinc finger-DNA complexes, and for designing and selecting Zif268 variants that will recognize other DNA sites.

Distinctive DNA conformation with enlarged major groove is found in Zn-finger-DNA and other protein-DNA complexes.

We have analyzed DNA conformations in a series of protein-DNA complexes, and we find that a distinctive conformation--with an enlarged major groove--occurs in a number of different complexes. During this analysis, we also developed a simplified model of DNA structure that illustrates the relative position of (i) the base pairs, (ii) the phosphate backbone, and (iii) the double-helical axis. This model highlights the key structural features of each duplex, facilitating the analysis and comparison of structures that are distinct from canonical A-DNA or B-DNA. Comparing DNA conformations in this way revealed that an otherwise unrelated set of protein-DNA complexes have interesting structural similarities, including an enlarged major groove. We refer to this class of structures as Beg-DNA (where eg means enlarged groove). Since related features occur in such a diverse set of protein-DNA complexes, we suggest that this conformation may have a significant role in protein-DNA recognition.