Hydrolysis of the RNA model substrate catalyzed by a binuclear ZrIV-substituted Keggin polyoxometalate

Abstract

The reactivity and solution behaviour of the binuclear ZrIV-substituted Keggin polyoxometalate (Et2NH2)8[{α-PW11O39Zr(μ-OH)(H2O)}2]·7H2O (ZrK 2 : 2) towards phosphoester bond hydrolysis of the RNA model substrate 2-hydroxypropyl-4-nitrophenyl phosphate (HPNP) was investigated at different reaction conditions (pD, temperature, concentration, and ionic strength). The hydrolysis of the phosphoester bond of HPNP, followed by means of 1H NMR spectroscopy, proceeded with an observed rate constant, kobs = 11.5(±0.42) × 10−5 s−1 at pD 6.4 and 50 °C, representing a 530-fold rate enhancement in comparison with the spontaneous hydrolysis of HPNP. 1H and 31P NMR spectra indicate that at these reaction conditions the only products of hydrolysis are p-nitrophenol and the corresponding cyclic phosphate ester. The pD dependence of kobs exhibits a bell-shaped profile, with the fastest rate observed at pD 6.4. The formation constant (Kf = 455 M−1) and catalytic rate constant (kc = 42 × 10−5 s−1) for the HPNP–ZrK 2 : 2 complex, activation energy (Ea) of 63.35 ± 1.82 kJ mol−1, enthalpy of activation (ΔH‡) of 60.60 ± 2.09 kJ mol−1, entropy of activation (ΔS‡) of −133.70 ± 6.13 J mol−1 K−1, and Gibbs activation energy (ΔG‡) of 102.05 ± 0.13 kJ mol−1 at 37 °C were calculated from kinetic experiments. Binding between ZrK 2 : 2 and the P–O bond of HPNP was evidenced by the change in the 31P chemical shift and signal linebroadening of the 31P atom in HPNP upon addition of ZrK 2 : 2. Based on 31P NMR experiments and isotope effect studies, a mechanism for HPNP hydrolysis in the presence of ZrK 2 : 2 was proposed.