The origin of the high stability of 3′-terminal uridine tetrads: contributions of hydrogen bonding, stacking interactions, and steric factors evaluated using modified oligonucleotide analogs

RNA G-quadruplexes fold almost exclusively into parallel-stranded structures and thus display much less structural diversity than their DNA counterparts. However, also among RNA G-quadruplexes peculiar structural elements can be found which are capable of reshaping the physico-chemical properties of the folded structure. A striking example is provided by a uridine tetrad (U-tetrad) placed on the 3′-terminus of the tetramolecular G-quadruplex. In this context, the U-tetrad adopts a unique conformation involving chain reversal and is responsible for a tremendous stabilization of the G-quadruplex (ΔT(m) up to 30°C). In this report, we attempt to rationalize the origin of this stabilizing effect by concurrent structural, thermal stability, and molecular dynamics studies of a series of G-quadruplexes with subtle chemical modifications at their 3′-termini. Our results provide detailed insights into the energetics of the “reversed” U-tetrad motif and the requirements for its formation. They point to the importance of the 2′OH to phosphate hydrogen bond and preferential stacking interactions for the formation propensity and stability of the motif.