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Nucleic Acids Research, 2003, Vol. 31, No. 17 5101-5107
© 2003 Oxford University Press

Mg2+-induced triplex formation of an equimolar mixture of poly(rA) and poly(rU)

Besik I. Kankia*

Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, 1479 Gortner Avenue, St Paul, MN 55108, USA

*Tel: +1 612 624 7468; Fax: +1 612 625 5780; Email: bkankia{at}umn.edu

Magnesium ions strongly influence the structure and biochemical activity of RNA. The interaction of Mg2+ with an equimolar mixture of poly(rA) and poly(rU) has been investigated by UV spectroscopy, isothermal titration calorimetry, ultrasound velocimetry and densimetry. Measurements in dilute aqueous solutions at 20°C revealed two differ ent processes: (i) Mg2+ binding to unfolded poly(rA)·poly(rU) up to [Mg2+]/[phosphate] = 0.25; and (ii) poly(rA)·2poly(rU) triplex formation at [Mg2+]/[phosphate] between 0.25 and 0.5. The enthalpies of these two different processes are favorable and similar to each other, ~–1.6 kcal mol–1 of base pairs. Volume and compressibility effects of the first process are positive, 8 cm3 mol–1 and 24 x 10–4 cm3 mol–1 bar–1, respectively, and correspond to the release of water molecules from the hydration shells of Mg2+ and the polynucleotides. The triplex formation is also accompanied by a positive change in compressibility, 14 x 10–4 cm3 mol–1 bar–1, but only a small change in volume, 1 cm3 mol–1. A phase diagram has been constructed from the melting experiments of poly(rA)·poly(rU) at a constant K+ concentration, 140 mM, and various amounts of Mg2+. Three discrete regions were observed, corresponding to single-, double- and triple-stranded complexes. The phase boundary corresponding to the transition between double and triple helical conformations lies near physiological salt concentrations and temperature.


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