Effect of Halogen Substitutions on dUMP to Stability
of Thymidylate Synthase/dUMP/mTHF Ternary Complex Using Molecular
Dynamics Simulation
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Abstract
The stability of the thymidylate
synthase (TS)/2-deoxyuridine-5-monophosphate
(dUMP)/5,10-methylene-5,6,7,8-tetrahydrofolate (mTHF) ternary complex
formation and Michael addition are considered as important steps that
are involved in the inhibition mechanism of the anticancer prodrug
5-fluorouracil (5-FU). Here, the effect of three different halogen
substitutions on the C-5 position of the dUMP (XdUMPs = FdUMP, CldUMP,
and BrdUMP), the normal substrate, on the stability of the TS/dUMP
and TS/dUMP/mTHF binary and ternary complexes, respectively, was investigated
via molecular dynamics simulation. The simulated results revealed
that the stability of all the systems was substantially increased
by mTHF binding to the catalytic pocket. In the ternary complex, a
much greater stabilization of the dUMP and XdUMPs through electrostatic
interactions, including charge–charge and hydrogen bond interactions,
was found compared to mTHF. An additional unique hydrogen bond between
the substituted fluorine of FdUMP and the hydroxyl group of the TS
Y94 residue was observed in both the binary and ternary complexes.
The distance between the S<sup>–</sup> atom of the TS C146
residue and the C6 atom of dUMP, at <4 Å in all systems, suggested
that a Michael addition with the formation of a S–C6 covalent
bond potentially occurred, although the hydrogen atom on C6 of dUMP
is substituted by a halogen atom. The MM/PBSA binding free energy
revealed the significant role of the bridging waters around the ligands
in the increased binding affinity (∼10 kcal/mol) of dUMP/XdUMP,
either alone or together with mTHF, toward TS. The order of the averaged
binding affinity in the ternary systems was found to be CldUMP ≈
FdUMP > dUMP > BrdUMP, suggesting that CldUMP could be a potent
candidate
TS inhibitor, the same as FdUMP (the metabolite form of 5-FU)