Force Field Design and Molecular Dynamics Simulations of the Carbapenem- and Cephamycin-Resistant Dinuclear Zinc Metallo-β-lactamase from <i>Bacteroides fragilis</i> and Its Complex with a Biphenyl Tetrazole Inhibitor

On the basis of molecular dynamics simulations, we investigate the dynamic properties of the carbapenem- and cephamycin-resistant dinuclear zinc metallo-β-lactamase from Bacteroides fragilis and its complex with a biphenyl tetrazole inhibitor, 2-butyl-6-hydroxy-3-[2‘-(1H-tetrazol-5-yl)biphenyl-4-ylmethyl]-3H-quinazolin-4-one 1 (L-159061). The results obtained with the newly developed force field parameters for the coordination environment of the catalytic zinc ions show that the active site gorge comprising major and minor loops gets deeper and narrower upon binding of the inhibitor, which supports the previous experimental implication that the structural flexibility of the loop structures plays a significant role in enzymatic action. In the presence of the inhibitor, the Trp32 side chain at the apex of the major loop covers the entrance of active site channel, thereby contributing to the stabilization of the enzyme−inhibitor complex. In addition to a direct coordination of the inhibitor tetrazole ring to the second zinc ion in the active site, the hydrogen bonding of Lys167 to the inhibitor carbonyl group and hydrophobic interactions between the inhibitor and side chains of loop residues prove to be significant binding forces of the enzyme−inhibitor complex

Role of Solvent Dynamics in Stabilizing the Transition State of RNA Hydrolysis by Hairpin Ribozyme

Structural and mechanistic studies of the hairpin ribozyme have been actively pursued over the last two decades to understand its catalytic strategy for RNA hydrolysis. Based on molecular dynamics simulations with the newly developed force field parameters for a vanadium−oxygen complex, we investigate the dynamic properties of the hairpin ribozyme in complex with a transition state analogue for the phosphodiester cleavage. The results indicate that the three nucleobases of the hairpin ribozyme (G8, A9, and A38) stabilize the negatively charged oxygen atoms in the transition state through the formation of five hydrogen bonds, which is consistent with the X-ray crystallographic data. In addition to the three catalytic nucleobases, several solvent molecules are also found to contribute to the catalytic action of the hairpin ribozyme by hydrogen bond stabilization of the negatively charged oxygens as well as by optimally positioning the catalytic nucleobases in the active site

MOESM1 of Accuracy enhancement in the estimation of molecular hydration free energies by implementing the intramolecular hydrogen bond effects

Additional file 1. Contains chemical structures, experimental and calculated solvation free energies of 763 molecules used in this study

Structural and Dynamical Basis of Broad Substrate Specificity, Catalytic Mechanism, and Inhibition of Cytochrome P450 3A4

Cytochrome P450 (CYP) 3A4 is responsible for the oxidative degradation of more than 50% of clinically used drugs. By means of molecular dynamics simulations with the newly developed force field parameters for the heme−thiolate group and its dioxygen adduct, we examine the differences in structural and dynamic properties between CYP3A4 in the resting form and its complexes with the substrate progesterone and the inhibitor metyrapone. The results indicate that the broad substrate specificity of CYP3A4 stems from the malleability of a loop (residues 211−218) that resides in the vicinity of the channel connecting the active site and bulk solvent. However, the high-amplitude motion of the flexible loop is found to be damped out upon binding of the inhibitor or the substrate in the active site. In the resting form of CYP3A4, a structural water molecule is bound to the sixth coordination position of the heme iron, stabilizing the octahedral coordination geometry. In addition to the direct coordination of metyrapone to the heme iron, the hydrogen bond interaction between the inhibitor carbonyl group and the side chain of Ser119 also contributes significantly to stabilizing the CYP3A4−metyrapone complex. On the other hand, progesterone is stabilized in the active site by the formation of two hydrogen bonds with Ser119 and Arg106, as well as by the van der Waals interactions with the heme and hydrophobic residues. The structural and dynamic features of the CYP3A4−progesterone complex indicate that the oxidative degradation of progesterone occurs through hydroxylation at the C16 position by the reactive oxygen coordinated to the heme iron

Computational Prediction of Molecular Hydration Entropy with Hybrid Scaled Particle Theory and Free-Energy Perturbation Method

Despite the importance of the knowledge of molecular hydration entropy (Δ<i>S</i>_hyd) in chemical and biological processes, the exact calculation of Δ<i><i>S</i></i>_hyd is very difficult, because of the complexity in solute–water interactions. Although free-energy perturbation (FEP) methods have been employed quite widely in the literature, the poor convergent behavior of the van der Waals interaction term in the potential function limited the accuracy and robustness. In this study, we propose a new method for estimating Δ<i><i>S</i></i>_hyd by means of combining the FEP approach and the scaled particle theory (or information theory) to separately calculate the electrostatic solute–water interaction term (Δ<i><i>S</i></i>_elec) and the hydrophobic contribution approximated by the cavity formation entropy (Δ<i><i>S</i></i>_cav), respectively. Decomposition of Δ<i><i>S</i></i>_hyd into Δ<i><i>S</i></i>_cav and Δ<i><i>S</i></i>_elec terms is found to be very effective with a substantial accuracy enhancement in Δ<i><i>S</i></i>_hyd estimation, when compared to the conventional full FEP calculations. Δ<i><i>S</i></i>_cav appears to dominate over Δ<i><i>S</i></i>_elec in magnitude, even in the case of polar solutes, implying that the major contribution to the entropic cost for hydration comes from the formation of a solvent-excluded volume. Our hybrid scaled particle theory and FEP method is thus found to enhance the accuracy of Δ<i><i>S</i></i>_hyd prediction by effectively complementing the conventional full FEP method

Discovery of Low Micromolar Dual Inhibitors for Wild Type and L1196M Mutant of Anaplastic Lymphoma Kinase through Structure-Based Virtual Screening

Although anaplastic lymphoma kinase (ALK) is involved in a variety of malignant human cancers, the emergence of constitutively active mutants with drug resistance has rendered it difficult to identify the new medicines for ALK-dependent cancers. To find the common inhibitors of the wild type ALK and the most abundant drug-resistant mutant (L1196M), we performed molecular docking-based virtual screening of a large chemical library in parallel for the two target proteins. As a consequence of augmenting the accuracy of the docking simulation by implementing a sophisticated hydration free energy term in the scoring function, 12 common inhibitors are discovered with the inhibitory activities ranging from submicromolar to low micromolar levels. The results of the binding free energy decomposition indicate that the biochemical potency of ALK inhibitors can be optimized by reducing the dehydration cost for binding to the receptor protein as well as by strengthening the interactions with amino acid residues in the ATP-binding site. The newly identified ALK inhibitors are found to have a little higher inhibitory activity for the L1196M mutant than for the wild type due to the strengthening of the hydrogen bond interactions in the ATP-binding site. Of the 12 common inhibitors, 2-(5-methyl-benzooxazol-2-ylamino)-quinazolin-4-ol (<b>3</b>) is anticipated to serve as a new molecular scaffold to optimize the biochemical potency because it exhibits low micromolar inhibitory activity with respect to both the wild type and L1196M mutant in spite of the low molecular weight (292.3 amu)

Application of Fragment-Based de Novo Design to the Discovery of Selective Picomolar Inhibitors of Glycogen Synthase Kinase‑3 Beta

A systematic fragment-based de novo design procedure was developed and applied to discover new potent and selective inhibitors of glycogen synthase kinase-3 beta (GSK3β). Candidate inhibitors were generated to simultaneously maximize the biochemical potency and the specificity for GSK3β through three design steps: identification of the optimal molecular fragments for the three sub-binding regions, design of proper linking moieties to connect the fragmental building blocks, and final scoring of the generated molecules. By virtue of modifying the ligand hydration free energy term in the scoring function using hybrid scaled particle theory and the extended solvent-contact model, we identified several GSK3β inhibitors with biochemical potencies ranging from low nanomolar to picomolar levels. Among them, the two most potent inhibitors (12 and 27) are anticipated to serve as promising starting points of drug discovery for various diseases caused by GSK3β because of the high specificity for the inhibition of GSK3β

Discovery of Picomolar ABL Kinase Inhibitors Equipotent for Wild Type and T315I Mutant via Structure-Based de Novo Design

Although the constitutively activated break-point cluster region–Abelson (ABL) tyrosine kinase is known to cause chronic myelogenous leukemia (CML), the prevalence of drug-resistant ABL mutants has made it difficult to develop effective anti-CML drugs. With the aim to identify new lead compounds for anti-CML drugs, we carried out a structure-based de novo design using the scoring function improved by implementing an accurate solvation free energy term. This approach led to the identification of ABL inhibitors equipotent for the wild type and the most drug-resistant T315I mutant of ABL at the picomolar level. Decomposition analysis of the binding free energy showed that a decrease in the desolvation cost for binding in the ATP-binding site could be as important as the strengthening of enzyme–inhibitor interaction to enhance the potency of an ABL inhibitor with structural modifications. A similar energetic feature was also observed in free energy perturbation (FEP) calculations. Consistent with the previous experimental and computational studies, the hydrogen bond interactions with the backbone groups of Met318 proved to be the most significant binding forces to stabilize the inhibitors in the ATP-binding sites of the wild type and T315I mutant. The results of molecular dynamics simulations indicated that the dynamic stabilities of the hydrogen bonds between the inhibitors and Met318 should also be considered in designing the potent common inhibitors of the wild-type and T315I mutant of ABL

Identification of Novel Inhibitors of Tropomyosin-Related Kinase A through the Structure-Based Virtual Screening with Homology-Modeled Protein Structure

Tropomyosin-related kinase A (TrkA) is a promising target for the development of cancer and pain therapeutics. Here, we report the first successful example of the use of a structure-based virtual screening to identify novel TrkA inhibitors. The accuracy of the virtual screening was improved by introducing an accurate solvation free energy term into the original AutoDock scoring function. We applied a drug design protocol involving homology modeling, docking analysis of a large chemical library, and enzyme inhibition assays to identify six structurally diverse TrkA inhibitors with <i>K</i>_<i>d</i> values ranging from 3 to 40 μM. The significant potencies and good physicochemical properties of these drug candidates strongly support their consideration in a development effort that would involve structure–activity relationship (SAR) studies to optimize the inhibitory activities. We also addressed the structural and energetic features associated with binding of the newly identified inhibitors in the ATP-binding site of TrkA. The results indicate that any structural modifications introduced for the purpose of enhancing the activity of TrkA inhibitors should maximize the attractive interactions within the ATP-binding site and simultaneously minimize the desolvation cost for complexation

Systematic Computational Design and Identification of Low Picomolar Inhibitors of Aurora Kinase A

Aurora kinase A (AKA) has served as an effective molecular target for the development of cancer therapeutics. A series of potent AKA inhibitors with the (4-methoxy-pyrimidin-2-yl)-phenyl-amine (MPPA) scaffold are identified using a systematic computer-aided drug design protocol involving structure-based virtual screening, de novo design, and free energy perturbation (FEP) simulations. To enhance the accuracy of the virtual screening to find a proper molecular core and de novo design to optimize biochemical potency, we preliminarily improved the scoring function by implementing a reliable hydration energy term. The overall design strategy proves successful to the extent that some inhibitors reveal exceptionally high potency at low picomolar levels; this was achieved by substituting phenyl, chlorine, and tetrazole moieties on the MPPA scaffold. The establishment of bidentate hydrogen bonds with backbone groups in the hinge region appears to be necessary for the high biochemical potency, consistent with the literature X-ray crystallographic data. The picomolar inhibitory activity also stems from the simultaneous formation of additional hydrogen bonds with the side chains of the hinge region and P-loop residues. The FEP simulation results show that the inhibitory activity surges to the low picomolar level because the interactions in the ATP-binding site of AKA become strong by structural modifications enough to overbalance the increase in dehydration cost. Because of the exceptionally high biochemical potency, the AKA inhibitors reported in this study are anticipated to serve as a new starting point for the discovery of anticancer medicine