QM/MM and SCRF studies of the ionization state of 8-methylpterin substrate bound to dihydrofolate reductase: existence of a low-barrier hydrogen bond.
Using combined semiempirical quantum mechanics and molecular mechanics (QM/MM) and ab initio self-consistent reaction field (SCRF) calculations, we determined that a low-barrier hydrogen bond (LBHB) is formed when the mechanism-based substrate 8-methylpterin binds to dihydrofolate reductase (DHFR). The substrate initially was assumed bound either in the ion-pair form corresponding to N3-protonated substrate hydrogen (H) bonded to the unprotonated (carboxylate) of the conserved Glu30 residue in the active site, or in the neutral-pair form corresponding to unprotonated substrate H bonded to the neutral (carboxylic acid) from of Glu30. The free energy of interaction of these H-bonded systems with the protein/solvent surroundings was computed using a coordinate-coupled free energy perturbation (FEP) method implemented within the molecular dynamics (MD) simulation scheme and using a semiempirical (PM3) QM/MM force field. The free energy obtained from the QM/MM force-field simulations corresponds most closely with the corresponding free energy component obtained from HF/6-31G* SCRF calculations using a value of 2 for the dielectric constant (epsilon) for the solvated protein. Calculations were performed at levels ranging from HF/6-31G to MP2/6-31G* to B3LYP/6-31 + G**, with varying dielectric constants. The energy-minimized path for motion of the proton in the H bond along a one-dimensional reaction coordinate was calculated at HF/6-31G, HF/6-31G* (epsilon = 1) and B3LYP/6-31G* (epsilon = 2) levels. These calculations identified a second neutral-pair complex, involving the 2-amino group of substrate, which also interacts with Glu30, which is lower in energy than the ion-pair form. A harmonic vibrational analysis shows that the first vibrational state appears to lie near or above the TS connecting potential energy minima corresponding to the two neutral-pair configurations, thus indicating an LBHB. Consequently, the H-bonded system will have a significant probability of being found in the ion-pair form, in agreement with experimental spectral studies indicating an enzyme-bound cation and suggesting that the LBHB would activate substrate towards hydride-ion transfer from NADPH.
Cummins
P L
PL
Computational Molecular Biology and Drug Design Group, John Curtin School of Medical Research, Australian National University, Canberra, Australia.
Gready
J E
JE
D000465:Algorithms; D002384:Catalysis; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D006860:Hydrogen Bonding; D007700:Kinetics; D015394:Molecular Structure; D011485:Protein Binding; D018892:Proton-Motive Force; D011622:Pterins / Q000737:chemistry* / Q000378:metabolism; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry* / Q000378:metabolism; D013816:Thermodynamics
D011622:Pterins; C080386:8-methylpterin; D013762:Tetrahydrofolate Dehydrogenase