Ground State Destabilization in Uracil DNA Glycosylase: Let's Not Forget "Tautomeric Strain" in Substrates

Abstract

Enzymes like uracil DNA glycosylase (UDG) can achieve ground state destabilization, by polarizing substrates to mimic rare tautomers. On the basis of computed nucleus independent chemical shifts, NICS(1)_zz, and harmonic oscillator model of electron delocalization (HOMED) analyses, of quantum mechanics (QM) and quantum mechanics/molecular mechanics (QM/MM) models of the UDG active site, uracil is strongly polarized when bound to UDG and resembles a tautomer >12 kcal/mol higher in energy. Natural resonance theory (NRT) analyses identified a dominant O2 imidate resonance form for residue bound 1-methyl-uracil. This "tautomeric strain" raises the energy of uracil, making uracilate a better than expected leaving group. Computed gas-phase S_N2 reactions of free and hydrogen bonded 1-methyl-uracil demonstrate the relationship between the degree of polarization in uracil and the leaving group ability of uracilate.

Figure 1.

(a) Proposed stepwise dissociative pathway for UDG. (b) Diketo (1a) and imidate (1b−1f) resonance forms of uracil.

Figure 2.

Computed tautomerization energies (ΔE_T) in the gas phase and at ε = 4 (values in parentheses), NICS(1)_zz, and HOMED for the canonical diketo form of uracil (1), and its keto−enol (2, 3) and dienol (4) tautomers.

Figure 3.

Truncated QM model of the UDG active site: Green dots and blue dashed lines indicate fixed positions of atoms and bond distances during geometry optimization. Computed NICS(1)_zz and HOMED values for uracil in the truncated QM model (values based on the QM/MM model are in parentheses).

Figure 4.

Relative energies (ΔE, in kcal/mol) of computed stationary points along the stepwise dissociative pathway of UDG, based on a constrained and truncated QM model (see the Computational Methods section for details) of the UDG active site. Values corrected for zero-point energy vibration (ZPVE) are shown in parentheses (ΔE_ZPVE, in kcal/mol). Geometries of all stationary points were optimized at ωB97X-D/6-31+G(d).