Kinetic hybrid i-motifs: Intercepting DNA with RNA to form a DNA2–RNA2 i-motif

We have constructed and characterized a long-lived hybrid DNA2–RNA2 i-motif that is kinetically formed by mixing equivalent amount of C-rich RNA (R) and C-rich DNA (D). Circular dichroism shows that these hybrids are distinct from their parent DNA4 or RNA4 i-motif. pH dependent CD and UV thermal melting experiments showed that the complexes were maximally stable at pH 4.5, the pKa of cytosine, consistent with the complex being held by CH+–C base pairs. Fluorescence studies confirmed their tetrameric nature and established the relative strand polarities of the RNA and DNA strands in the complex. These showed that in a hybrid D2R2 i-motif two DNA strands occupy one narrow groove and the two RNA strands occupy the other. This suggests that even the sugar–sugar interactions are highly specific. Interestingly, this hybrid slowly disproportionates into DNA4 i-motifs and ssRNA which would be valuable to study intermediates in DNA4 i-motif formation.