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Slow-Onset Inhibition of Mycobacterium tuberculosis InhA: Revealing Molecular Determinants of Residence Time by MD Simulations.

Merget B, Sotriffer CA - PLoS ONE (2015)

Bottom Line: Whereas the diphenyl ether inhibitors 6PP and triclosan (TCL) do not show loop ordering and thus, no slow-binding inhibition and high koff values, the slightly modified PT70 leads to an ordered loop and a residence time of 24 minutes.The individual simulations show comparable conformational features with respect to both the binding pocket and the SBL, allowing to define five recurring conformational families.The most abundant conformation besides the stable EI* state is characterized by a shift of Ile202 and Val203 toward the hydrophobic pocket of InhA.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pharmacy and Food Chemistry, University of Würzburg, Am Hubland, D-97074, Würzburg, Germany.

ABSTRACT
An important kinetic parameter for drug efficacy is the residence time of a compound at a drug target, which is related to the dissociation rate constant koff. For the essential antimycobacterial target InhA, this parameter is most likely governed by the ordering of the flexible substrate binding loop (SBL). Whereas the diphenyl ether inhibitors 6PP and triclosan (TCL) do not show loop ordering and thus, no slow-binding inhibition and high koff values, the slightly modified PT70 leads to an ordered loop and a residence time of 24 minutes. To assess the structural differences of the complexes from a dynamic point of view, molecular dynamics (MD) simulations with a total sampling time of 3.0 µs were performed for three ligand-bound and two ligand-free (perturbed) InhA systems. The individual simulations show comparable conformational features with respect to both the binding pocket and the SBL, allowing to define five recurring conformational families. Based on their different occurrence frequencies in the simulated systems, the conformational preferences could be linked to structural differences of the respective ligands to reveal important determinants of residence time. The most abundant conformation besides the stable EI* state is characterized by a shift of Ile202 and Val203 toward the hydrophobic pocket of InhA. The analyses revealed potential directions for avoiding this conformational change and, thus, hindering rapid dissociation: (1) an anchor group in 2'-position of the B-ring for scaffold stabilization, (2) proper occupation of the hydrophobic pocket, and (3) the introduction of a barricade substituent in 5'-position of the diphenyl ether B-ring.

No MeSH data available.


Related in: MedlinePlus

2D density plot for the ether dihedral angles α and β of the unbound ligands PT70 (left) and 6PP (right) based on a 150 ns MD simulation in aqueous solution.The dihedral angles α (COH-C-O-C) and β (C-O-C-CMe/H) are illustrated in Fig 2.
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pone.0127009.g010: 2D density plot for the ether dihedral angles α and β of the unbound ligands PT70 (left) and 6PP (right) based on a 150 ns MD simulation in aqueous solution.The dihedral angles α (COH-C-O-C) and β (C-O-C-CMe/H) are illustrated in Fig 2.

Mentions: While 6PP is a rapid reversible inhibitor, PT70 binds with a residence time of 24 minutes (Fig 2), although the ortho-methyl group at the B-ring of PT70 is the only structural difference [7, 13]. Interestingly, for the 6PP complex the simulations indicate a reduced stability of the Family 1 state in comparison to the PT70 complex, and the conformational Families 2 and 3 are significantly more frequent in the 6PP simulation. With the ortho-methyl moiety as the only substitution, this difference appears as the logical origin for these observations. To investigate the effect of ortho-methyl substitution on the ligand conformations (which are mainly determined by the torsions around the two ether bonds), two additional 150 ns MD simulations were conducted for each ligand solvated in a water box. By measuring the dihedral angles of the ether moiety along the trajectory, a 2D density map of the (C-O-C-CMe/H)-dihedral βversus the (COH-C-O-C)-dihedral α was generated for each ligand (Fig 10). The strong peaks in the distribution of the PT70 angle pairs suggest that fewer conformations are populated compared to 6PP. Hence, as expected, the ortho-substituted PT70 is more constrained in its intramolecular mobility, hindering the Ile202 movement toward the hydrophobic pocket to a greater extent than the unsubstituted 6PP. This very likely accounts for the enhanced occurrence of Families 2 and 3 in the case of 6PP and for the (on average) larger RMS deviations and fluctuations of the ligand in the binding pocket (Table 2). Interestingly, also the hexyl chain of 6PP shows higher mobility than the PT70 hexyl chain in the binding pocket (Table 2, Supporting Information S6 Fig). In summary, the conformational stabilization of PT70 by the ortho-methyl group appears to translate directly to increased SBL stabilization and retention of a Family 1 conformation.


Slow-Onset Inhibition of Mycobacterium tuberculosis InhA: Revealing Molecular Determinants of Residence Time by MD Simulations.

Merget B, Sotriffer CA - PLoS ONE (2015)

2D density plot for the ether dihedral angles α and β of the unbound ligands PT70 (left) and 6PP (right) based on a 150 ns MD simulation in aqueous solution.The dihedral angles α (COH-C-O-C) and β (C-O-C-CMe/H) are illustrated in Fig 2.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4440617&req=5

pone.0127009.g010: 2D density plot for the ether dihedral angles α and β of the unbound ligands PT70 (left) and 6PP (right) based on a 150 ns MD simulation in aqueous solution.The dihedral angles α (COH-C-O-C) and β (C-O-C-CMe/H) are illustrated in Fig 2.
Mentions: While 6PP is a rapid reversible inhibitor, PT70 binds with a residence time of 24 minutes (Fig 2), although the ortho-methyl group at the B-ring of PT70 is the only structural difference [7, 13]. Interestingly, for the 6PP complex the simulations indicate a reduced stability of the Family 1 state in comparison to the PT70 complex, and the conformational Families 2 and 3 are significantly more frequent in the 6PP simulation. With the ortho-methyl moiety as the only substitution, this difference appears as the logical origin for these observations. To investigate the effect of ortho-methyl substitution on the ligand conformations (which are mainly determined by the torsions around the two ether bonds), two additional 150 ns MD simulations were conducted for each ligand solvated in a water box. By measuring the dihedral angles of the ether moiety along the trajectory, a 2D density map of the (C-O-C-CMe/H)-dihedral βversus the (COH-C-O-C)-dihedral α was generated for each ligand (Fig 10). The strong peaks in the distribution of the PT70 angle pairs suggest that fewer conformations are populated compared to 6PP. Hence, as expected, the ortho-substituted PT70 is more constrained in its intramolecular mobility, hindering the Ile202 movement toward the hydrophobic pocket to a greater extent than the unsubstituted 6PP. This very likely accounts for the enhanced occurrence of Families 2 and 3 in the case of 6PP and for the (on average) larger RMS deviations and fluctuations of the ligand in the binding pocket (Table 2). Interestingly, also the hexyl chain of 6PP shows higher mobility than the PT70 hexyl chain in the binding pocket (Table 2, Supporting Information S6 Fig). In summary, the conformational stabilization of PT70 by the ortho-methyl group appears to translate directly to increased SBL stabilization and retention of a Family 1 conformation.

Bottom Line: Whereas the diphenyl ether inhibitors 6PP and triclosan (TCL) do not show loop ordering and thus, no slow-binding inhibition and high koff values, the slightly modified PT70 leads to an ordered loop and a residence time of 24 minutes.The individual simulations show comparable conformational features with respect to both the binding pocket and the SBL, allowing to define five recurring conformational families.The most abundant conformation besides the stable EI* state is characterized by a shift of Ile202 and Val203 toward the hydrophobic pocket of InhA.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pharmacy and Food Chemistry, University of Würzburg, Am Hubland, D-97074, Würzburg, Germany.

ABSTRACT
An important kinetic parameter for drug efficacy is the residence time of a compound at a drug target, which is related to the dissociation rate constant koff. For the essential antimycobacterial target InhA, this parameter is most likely governed by the ordering of the flexible substrate binding loop (SBL). Whereas the diphenyl ether inhibitors 6PP and triclosan (TCL) do not show loop ordering and thus, no slow-binding inhibition and high koff values, the slightly modified PT70 leads to an ordered loop and a residence time of 24 minutes. To assess the structural differences of the complexes from a dynamic point of view, molecular dynamics (MD) simulations with a total sampling time of 3.0 µs were performed for three ligand-bound and two ligand-free (perturbed) InhA systems. The individual simulations show comparable conformational features with respect to both the binding pocket and the SBL, allowing to define five recurring conformational families. Based on their different occurrence frequencies in the simulated systems, the conformational preferences could be linked to structural differences of the respective ligands to reveal important determinants of residence time. The most abundant conformation besides the stable EI* state is characterized by a shift of Ile202 and Val203 toward the hydrophobic pocket of InhA. The analyses revealed potential directions for avoiding this conformational change and, thus, hindering rapid dissociation: (1) an anchor group in 2'-position of the B-ring for scaffold stabilization, (2) proper occupation of the hydrophobic pocket, and (3) the introduction of a barricade substituent in 5'-position of the diphenyl ether B-ring.

No MeSH data available.


Related in: MedlinePlus