Effect of Halogen Substitutions on dUMP to Stability of Thymidylate Synthase/dUMP/mTHF Ternary Complex Using Molecular Dynamics Simulation

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(-) 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).

Metadynamics supports molecular dynamics simulation-based binding affinities of eucalyptol and beta-cyclodextrin inclusion complexes

“…59 bCD and its dimethyl and hydroxypropyl derivatives were treated using the Glycam-06h carbohydrate force eld. 60 The standard procedures based on our previous studies for small organic molecules [61][62][63][64] were used for the preparation of the partial atomic charges and parameters of eucalyptol. Briey, the optimized structure of eucalyptol was used to calculate the electrostatic potential (ESP) charges with the HF/6-31G(d) level of theory using Gaussian 09.…”

Expression of Folate Pathway Genes in Stage III Colorectal Cancer Correlates with Recurrence Status Following Adjuvant Bolus 5-FU-Based Chemotherapy

“…LV is considered to increase the concentration of the natural reduced folate cofactor [6R]-5,10-methylenetetrahydrofolate (methyleneTHF) in the cells (2). This cofactor forms a ternary complex with the enzyme thymidylate synthase (TS) in a reaction in which deoxyuridine monophosphate (dUMP) is transformed to deoxythymidine monophosphate (dTMP) (3). After administration with 5-FU, a fluorinated form of dUMP is formed intracellularly, which leads to inhibition of the TS enzyme.…”

Simultaneous, But Not Consecutive, Combination With Folinate Salts Potentiates 5-Fluorouracil Antitumor Activity In Vitro and In Vivo

“…Instead, and surprisingly, the sequential combination of NaLV or CaLV (1 h before) and then 5-FU resulted not additive but even antagonistic in a 24 and 72h treatment. Moreover, the simultaneous combination (but not the sequential one) of 5-FU and NaLV or CaLV encodes the 5-FU target thymidylate synthase, is known to affect response to treatment 33 . In our experiments, the simultaneous combination (but not the sequential one) of 5-FU and NaLV or CaLV (although in a lesser extent) significantly inhibited the TYMS gene expression after 24 h exposure.…”