Effect of Sequence on the Conformation of DNA Holliday Junctions

Abstract

Structures of the DNA sequences d(CCGGCGCCGG) and d(CCAGTACbr(5)UGG) are presented here as four-way Holliday junctions in their compact stacked-X forms, with antiparallel alignment of the DNA strands. Thus, the ACC-trinucleotide motif, previously identified as important for stabilizing the junction, is now extended to PuCPy, where Pu is either an adenine or guanine, and Py is either a cytosine, 5-methylcytosine, or 5-bromouracil but not thymine nucleotide. We see that both sequence and base substituents affect the geometry of the junction in terms of the interduplex angle as well as the previously defined conformational variables, J(roll) (the rotation of the stacked duplexes about their respective helical axis) and J(slide) (the translational displacement of the stacked duplexes along their respective helical axis). The structures of the GCC and parent ACC containing junctions fall into a distinct conformational class that is relatively undistorted in terms of J(slide) and J(roll), with interduplex angles of 40-43 degrees. The substituted ACbr(5)U structure, however, is more akin to that of the distorted methylated ACm(5)C containing junction, with J(slide) (>or=2.3 A) and a similar J(roll) (164 degrees) opening the major groove-side of the junction, but shows a reduced interduplex angle. In contrast, the analogous d(CCAGTACTGG) sequence has to date been crystallized only as resolved B-DNA duplexes. This suggests that there is an electronic effect of substituents at the pyrimidine Py position on the stability of four-stranded junctions. The single-crystal structures presented here, therefore, show how sequence affects the detailed geometry, and subsequently, the associated stability and conformational dynamics of the Holliday junction.

How sequence defines structure: A crystallographic map of DNA structure and conformation

“…3). We note, however, that this interaction can be partially replaced by an analogous Br⅐⅐⅐O halogen bond, as in the structure of the ACbr 5 U junction (18,28).…”

“…These solutions are similar to previous conditions used to crystallize DNA oligomers in this laboratory (18). Each sequence was subjected to the full range of CaCl 2 and spermine concentrations, with each crystal form refined individually to obtain diffraction-quality single crystals.…”

Halogen bonds in biological molecules

“…wo recent biomolecular single-crystal structures, a fourstranded DNA Holliday junction (1) and an ultrahighresolution structure (0.66 Å) of the enzyme aldose reductase complex with a halogenated inhibitor (2), revealed unusually short Br⅐⅐⅐O contacts [Ϸ3.0 Å, or Ϸ12% shorter than the sum of their van der Waals radii (R vdW )]. The atypical contact in the enzyme complex was attributed to an electrostatic interaction between the polarized bromine and the lone pair electrons of the oxygen atom of a neighboring threonine side chain (3).…”

Influence of C-5 halogenation of uridines on hairpin versus duplex RNA folding

“…An even more dramatic structural effect was observed for the bromination of the DNA sequence d(CCAGTACTGG). This sequence folds as a duplex, but it folds as a four-way junction when a bromo-uridine is substituted for the second thymine in the sequence (Hays et al 2003). In this situation, analyzed in detail by Auffinger et al (2004), the bromine atom, being involved in a crucial halogen bond (z3.0 Å distance) with an adjacent phosphate oxygen, appears to be responsible for the conformational change.…”