DNA bending by an adenine–thymine tract and its role in gene regulation

Abstract

To gain insight into the structural basis of DNA bending by adenine-thymine tracts (A-tracts) and their role in DNA recognition by gene-regulatory proteins, we have determined the crystal structure of the high-affinity DNA target of the cancer-associated human papillomavirus E2 protein. The three independent B-DNA molecules of the crystal structure determined at 2.2-Å resolution are examples of A-tract-containing helices where the global direction and magnitude of curvature are in accord with solution data, thereby providing insights, at the base pair level, into the mechanism of DNA bending by such sequence motifs. A comparative analysis of E2-DNA conformations with respect to other structural and biochemical studies demonstrates that (i) the A-tract structure of the core region, which is not contacted by the protein, is critical for the formation of the high-affinity sequence-specific protein-DNA complex, and (ii) differential binding affinity is regulated by the intrinsic structure and deformability encoded in the base sequence of the DNA target.A-tract ͉ papillomavirus E2-DNA target ͉ transcriptional control T he relationship between DNA sequence, structure, and function has been studied and discussed extensively for the last 20 years. A particular effort has been directed toward the structural elucidation of short runs of four to six adeninethymine residues, known as A-tracts, in an attempt to reveal the structural basis of DNA curvature induced by such sequence motifs when they are inserted in phase with the helical periodicity (1, 2). Despite numerous efforts, including x-ray crystallographic and solution studies, the structural basis of A-tractinduced bending has remained enigmatic (3). Because no single structure could explain the whole phenomenon, it was necessary to rely on models, several of which had been proposed (4). They generally conform to the gel migration data, which suggest that the center of curvature is toward the minor groove of the A-tracts and toward the major groove of the intervening general sequences (5). However, they differ substantially in the details of the stereochemical origin of curvature. This issue is of particular biological significance, as sequence-dependent DNA curvature or bending is an important determinant of DNA recognition by proteins (6).The E2 regulatory system of human papillomaviruses provides an example where sequence-specific binding of proteins to A-tracts is crucial to the organism's life cycle. The E2 proteins from all viral strains activate or repress transcription in a context-dependent manner and are required for the initiation of replication in vivo. Their function depends on sequence-specific binding to a highly conserved 12-bp sequence of the general form ACCGNNNNCGGT, where N4 is variable (7,8). However, the E2 binding sites in the human papillomavirus (HPV) genomes, including the cancer-related strains HPV-16 and HPV-18 (9), exhibit a further level of specificity in the interaction with their cognate E2 proteins. This specificity is manifested by th...

Structural origins of adenine-tract bending

“…Bifurcated hydrogen bonds, however, are not necessary for A-tract curvature (34). Buckle may play an important role in bending of A-tracts and has been observed before in AATT tracts (35). We observe a significant change (12°at the 5Ј end and 25°at the 3Ј end) of buckling from one base pair to the next at the junctions (Fig.…”

Right-handed double-helix ultrashort DNA yields chiral nematic phases with both right- and left-handed director twist

“…However, a few clues suggest that the structural variability among sequences can play a significant role. The reported structure of DD has an appreciable bend of the helical axis (19), which is even larger for the ACC structure (30), indicating that oligomeric duplexes are not necessarily straight. Both sequences are RH whereas their concentration at the isotropic-nematic transition (c IN ) is rather high when compared to the whole set of data in Table 1.…”

Transition state for protein–DNA recognition

“…The free BOV oligonucleotide adopts a nearly straight conformation, whereas a sequence similar to the wildtype oligonucleotide is prebent toward the minor groove in the absence of E2C, although further bending takes place on binding (30). The mutated linker destabilizes the final complex by 1.4 kcal/mol relative to the wild-type linker, as expected for a reduced bendability.…”

“…The four linker bases bend on binding, whereas the two conserved half-sites do not significantly change conformation (9,14,22,23,30). Because the linker bases are not in direct contact with the protein in the E2C-DNA complex, but their composition does affect the binding energetics (23,30), the energetic contribution of DNA deformation is called ''indirect readout'' (12)(13)(14)(15).…”

“…On binding of HPV16 E2C to DNA, the conformational changes are relatively large for the DNA and small for the protein (9,14,23,24,29,30). However, the DNA recognition elements of E2C, helix1, and the ␤2-␤3 loop, show substantial flexibility in the unbound form (24,31).…”