Structure of three-way DNA junctions. 2. Effects of extra bases and mismatches.

The structure of three-way DNA junctions, containing two linear double helices (arms) and a hairpin as a third arm, was studied by means of a cyclization technique. In addition to branched molecules containing perfect base-pairing in helical parts, three-way junctions with mismatches and extra non-complementary nucleotides (bulges) at junction points were studied. Molecules thus designed were ligated at identical conditions and their geometry was compared through the analysis of the efficiency of circle formation. The analysis showed that irregularities in base pairing listed above dramatically change the static and dynamic structural characteristics of the three-way junctions. All mismatches facilitate the kink between linear arms, but quantitatively, the effect depends on the position of the mismatch. The effect is maximal for GG-mismatch placed at the hairpin junction point. The results for bulges are of different kind, and they lead us to conclude that the three-way DNA junction with unpaired nucleotides adopts a T-like geometry with an angle around 90 degrees between arms containing the bulge and two other arms coaxially stacked. Broad distribution of circles indicates that this T-form geometry of bulge-containing junction is more flexible than initial pyramidal structure predominantly due to high mobility of the third arm.