Diversity in DNA recognition by p53 revealed by crystal structures with Hoogsteen base pairs

Abstract

p53 binds as a tetramer to DNA targets consisting of two decameric half-sites separated by a variable spacer. Here we present high-resolution crystal structures of complexes between p53 core-domain tetramers and DNA targets consisting of contiguous half-sites. In contrast to previously reported p53-DNA complexes that display standard Watson-Crick base pairs, the newly reported structures exhibit non-canonical Hoogsteen base-pairing geometry at the central A/T doublet of each half-site. Structural and computational analyses demonstrate that the Hoogsteen geometry distinctly modulates the B-DNA helix in terms of local shape and electrostatic potential which together with the contiguous DNA configuration results in enhanced protein-DNA and protein-protein interactions compared to non-contiguous half-sites. Our results suggest a mechanism, which relates spacer length to protein-DNA binding affinity. Our findings also expand the current understanding of protein-DNA recognition and establish the structural and chemical properties of Hoogsteen base pairs as the basis for a novel mode of sequence readout.

Electron microscopy studies on the quaternary structure of p53 reveal different binding modes for p53 tetramers in complex with DNA

“…We fitted the atomic structures for DBD tetramers on DNA as rigid bodies inside our EM map for class III. While the structures on continuous DNA (20)(21)(22) are not compatible with the current architecture, the DBD tetramer bound to discontinuous DNA (19) shows a good agreement with our envelope (Fig. 3E).…”

“…Only 25% of the population displays a compact DBD tetramer (class III) where dimers appear rotated relative to each other. This architecture is compatible with the atomic structures of DBD engaged in specific DNA recognition of discontinuous DNA (19,22) but cannot accommodate structures of DBD bound to continuous half-sites that display dimers arranged in parallel fashion (20)(21)(22). Our DNA probe, however, contains continuous half-sites and should be overtwisted to accommodate the four DBD in specific binding.…”

“…The Tet domain forms a dimer of dimers (16)(17)(18). Isolated DBDs self assemble on DNA to form tetramers that have been described bound to two half-site DNA duplexes (19), in oligomers stabilized by crosslinking between DBD and cognate DNA (20), and bound to full consensus sites without chemical modification (21,22). These structures show varying dimer-dimer contacts and the overall assembly of the core domains on DNA is radically different from the one described in the first DBD · DNA complex solved (23).…”

Domain–domain interactions in full-length p53 and a specific DNA complex probed by methyl NMR spectroscopy

“…The methyl NMR spectrum corroborated that change: The signal of M243 split into two broad cross-peaks, and the "monomeric" cross-peak that is prevalent in free p53 spectra disappeared completely (one M243 signal was overlayed with the "dimeric" M243 from free p53 spectra and the other was downfield-shifted). Although disappearance of the "monomeric" signal indicated that all four p53C domains interacted with DNA, the splitting pattern of M243 was difficult to interpret in the light of X-ray studies of p53 complexes with 20-bp nucleotides (31,32). Symmetry of the crystal lattice in those structures implies chemical equivalence of M243 and L3 loops in the DNA complexes with contiguous p53-binding sites (Fig.…”

DNA breathes Hoogsteen

“…They also play an important role in sequence-specific targeting duplex DNA with PNA 4 and they have been found to be formed in complexes of duplex DNA with some proteins. 5 However, the spontaneous formation of HG pairs within regular B-DNA under normal conditions would seem highly unlikely due to several reasons:…”