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. 2017 Nov 24;22(12):2055.
doi: 10.3390/molecules22122055.

Species-Specific Inactivation of Triosephosphate Isomerase from Trypanosoma brucei: Kinetic and Molecular Dynamics Studies

Alejandra Vázquez-Raygoza  1 , Lucia Cano-González  2 , Israel Velázquez-Martínez  2 , Pedro Josué Trejo-Soto  2 , Rafael Castillo  2 , Alicia Hernández-Campos  2 , Francisco Hernández-Luis  2 , Jesús Oria-Hernández  3 , Adriana Castillo-Villanueva  3 , Claudia Avitia-Domínguez  1 , Erick Sierra-Campos  4 , Mónica Valdez-Solana  4 , Alfredo Téllez-Valencia  1
Affiliations

Species-Specific Inactivation of Triosephosphate Isomerase from Trypanosoma brucei: Kinetic and Molecular Dynamics Studies

Alejandra Vázquez-Raygoza et al. Molecules. .

Abstract

Human African Trypanosomiasis (HAT), a disease that provokes 2184 new cases a year in Sub-Saharan Africa, is caused by Trypanosoma brucei. Current treatments are limited, highly toxic, and parasite strains resistant to them are emerging. Therefore, there is an urgency to find new drugs against HAT. In this context, T. brucei depends on glycolysis as the unique source for ATP supply; therefore, the enzyme triosephosphate isomerase (TIM) is an attractive target for drug design. In the present work, three new benzimidazole derivatives were found as TbTIM inactivators (compounds 1, 2 and 3) with an I50 value of 84, 82 and 73 µM, respectively. Kinetic analyses indicated that the three molecules were selective when tested against human TIM (HsTIM) activity. Additionally, to study their binding mode in TbTIM, we performed a 100 ns molecular dynamics simulation of TbTIM-inactivator complexes. Simulations showed that the binding of compounds disturbs the structure of the protein, affecting the conformations of important domains such as loop 6 and loop 8. In addition, the physicochemical and drug-like parameters showed by the three compounds suggest a good oral absorption. In conclusion, these molecules will serve as a guide to design more potent inactivators that could be used to obtain new drugs against HAT.

Keywords: enzymatic kinetics; human African trypanosomiasis; molecular dynamics simulation; triosephosphate isomerase.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Structure of TbTIM inactivators.
Figure 1
Figure 1
Structure of TbTIM inactivators.
Figure 2
Figure 2
Activity of TbTIM (left panel) and plots of the pseudo-first-order rate constants (right panel) at different concentrations of compounds (a) 1; (b) 2 and (c) 3. I50 value was defined as the concentration of compound needed to reduce the enzymatic activity to 50% and determined through curves at different compound concentrations and a Hill coefficient, n, is a measure of the degree of cooperativity of the ligands.
Figure 2
Figure 2
Activity of TbTIM (left panel) and plots of the pseudo-first-order rate constants (right panel) at different concentrations of compounds (a) 1; (b) 2 and (c) 3. I50 value was defined as the concentration of compound needed to reduce the enzymatic activity to 50% and determined through curves at different compound concentrations and a Hill coefficient, n, is a measure of the degree of cooperativity of the ligands.
Figure 3
Figure 3
Effect of compounds 1, 2 and 3 at different concentrations of TbTIM.
Figure 4
Figure 4
Root mean square deviation (RMSD) of free enzyme (Apo-TbTIM) and the TbTIM-compound complexes.
Figure 5
Figure 5
(a) Residue Mean-Square Fluctuations from monomer A and B (* p < 0.05). Superimposed average structures of Apo-TbTIM (blue) with TbTIM-1 (b); TbTIM-2 (c); and TbTIM-3 (d). The alignments were made based on backbone.
Figure 6
Figure 6
Binding mode of compounds 1, 2 and 3 on TbTIM (Light Gray ribbons monomer A and Light turquoise ribbons monomer B. (a) two molecules of compound 1 (blue sticks); (b) two molecules of compound 2 (red sticks); and (c) compound 3 (yellow sticks). H-bonds are depicted as dotted lines.
Figure 7
Figure 7
Catalytic loop 6 dynamics in (a) Apo-TbTIM; (b) TbTIM-1; (c) TbTIM-2; and (d) TbTIM-3. Cluster 1 (gray), Cluster 2 (cyan), Cluster 3 (magenta), Cluster 4 (yellow) and Cluster 5 (pink). The alignments were made based on backbone.
Figure 7
Figure 7
Catalytic loop 6 dynamics in (a) Apo-TbTIM; (b) TbTIM-1; (c) TbTIM-2; and (d) TbTIM-3. Cluster 1 (gray), Cluster 2 (cyan), Cluster 3 (magenta), Cluster 4 (yellow) and Cluster 5 (pink). The alignments were made based on backbone.
Figure 8
Figure 8
Loop 8 dynamics in (a) Apo-TbTIM; (b) TbTIM-1; (c) TbTIM-2; and (d) TbTIM-3. Cluster 1 (gray), Cluster 2 (cyan), Cluster 3 (magenta), Cluster 4 (yellow) and Cluster 5 (pink). The alignments were made based on backbone.
Figure 9
Figure 9
Movement of side chains from catalytic site residues in the Apo-TbTIM (blue sticks) and in complex with (a) compound 1; (b) compound 2; and (c) compound 3.
Figure 10
Figure 10
Inactivation of HsTIM by compounds 1 (squares), 2 (circles) and 3 (triangles).
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