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

Abstract

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 degrees 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, approximately -1.6 kcal x mol(-1) of base pairs. Volume and compressibility effects of the first process are positive, 8 cm3 x mol(-1) and 24 x 10(-4) cm3 x mol(-1) x 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 x mol(-1) x bar(-1), but only a small change in volume, 1 cm3 x 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.