Yurie Watanabe

Evaluation of Influence of Single Nucleotide Polymorphisms in Cytochrome P450 2B6 on Substrate Recognition Using Computational Docking and Molecular Dynamics Simulation

In this study, we investigated the influence of single nucleotide polymorphisms on the conformation of mutated cytochrome P450 (CYP) 2B6 proteins using molecular dynamics (MD) simulation. Some of these mutations influence drug metabolism activities, leading to individual variations in drug efficacy and pharmacokinetics. Using computational docking, we predicted the structure of the complex between the antimalarial agent artemether and CYP2B6 whose conformations were obtained by MD simulation. The simulation demonstrated that the entire structure of the protein changes even when a single residue is mutated. Moreover, the structural flexibility is affected by the mutations and it may influence the enzyme activity. The results suggest that some of the inactive mutants cannot recognize artemether due to structural changes caused by the mutation.

Molecular Dynamics Simulations to Investigate the Influences of Amino Acid Mutations on Protein Three-Dimensional Structures of Cytochrome P450 2D6.1, 2, 10, 14A, 51, and 62

Many natural mutants of the drug metabolizing enzyme cytochrome P450 (CYP) 2D6 have been reported. Because the enzymatic activities of many mutants are different from that of the wild type, the genetic polymorphism of CYP2D6 plays an important role in drug metabolism. In this study, the molecular dynamics simulations of the wild type and mutants of CYP2D6, CYP2D6.1, 2, 10, 14A, 51, and 62 were performed, and the predictions of static and dynamic structures within them were conducted. In the mutant CYP2D6.10, 14A, and 61, dynamic properties of the F-G loop, which is one of the components of the active site access channel of CYP2D6, were different from that of the wild type. The F-G loop acted as the “hatch” of the channel, which was closed in those mutants. The structure of CYP2D6.51 was not converged by the simulation, which indicated that the three-dimensional structure of CYP2D6.51 was largely different from that of the wild type. In addition, the intramolecular interaction network of CYP2D6.10, 14A, and 61 was different from that of the wild type, and it is considered that these structural changes are the reason for the decrease or loss of enzymatic activities. On the other hand, the static and dynamic properties of CYP2D6.2, whose activity was normal, were not considerably different from those of the wild type.

Prediction of three-dimensional structures and structural flexibilities of wild-type and mutant cytochrome P450 1A2 using molecular dynamics simulations.

In this study, we investigated the effect of genetic polymorphism on the three-dimensional (3D) conformation of cytochrome P450 1A2 (CYP1A2) using molecular dynamics (MD) simulations. CYP1A2, a major drug-metabolizing enzyme among cytochrome P450 enzymes (CYPs), is known to have many variant alleles. The genetic polymorphism of CYP1A2 may cause individual differences in the pharmacokinetics of medicines. By performing 100ns or longer MD simulations, we investigated the influence of amino acid mutation on the 3D structures and the dynamic properties of proteins. The results show that the static structures were changed by the mutations of amino acid residues, not only near the mutated residues but also in distant portions of the proteins. Moreover, the mutation of only one amino acid was shown to change the structural flexibility of proteins, which may influence the substrate recognition and enzymatic activity. Our results clearly suggest that it is necessary to investigate the dynamic property as well as the static 3D structure for understanding the change of the enzymatic activity of mutant CYP1A2.

Effects of initial settings on computational protein–ligand docking accuracies for several docking programs

In this study, the influences of initial settings, i.e. initial conformations, configurations and docking parameters, on docking results were investigated. The conformations used in the study were generated by the CAMDAS program. After the conformational search calculations, five structures were selected from the conformer groups according to their conformation energies and root mean square deviations against crystal structures; for example, the lowest energy conformer, as well as the closest and farthest conformers to the crystal structure, was retrieved. Several docking parameter settings were used (default, high speed, generating 50 poses). In this study, docking calculations were conducted using the GOLD, eHiTS, AutoDock, AutoDock vina, FRED and DOCK programs. The success rates of GOLD, eHiTS and FRED were better than those of AutoDock, AutoDock vina and DOCK. The docking results using the farthest conformations were worse than those obtained using other conformations, indicating that some conformation search for the ligand molecule should be performed before the docking calculations.

Predicting the structures of complexes between phosphoinositide 3-kinase (PI3K) and romidepsin-related compounds for the drug design of PI3K/histone deacetylase dual inhibitors using computational docking and the ligand-based drug design approach.

Predictions of the three-dimensional (3D) structures of the complexes between phosphoinositide 3-kinase (PI3K) and two inhibitors were conducted using computational docking and the ligand-based drug design approach. The obtained structures were refined by structural optimizations and molecular dynamics (MD) simulations. The ligands were located deep inside the ligand binding pocket of the p110α subunit of PI3K, and the hydrogen bond formations and hydrophobic effects of the surrounding amino acids were predicted. Although rough structures were obtained for the PI3K-inhibitor complexes before the MD simulations, the refinement of the structures by these simulations clarified the hydrogen bonding patterns of the complexes.

Investigation of substrate recognition for cytochrome P450 1A2 mediated by water molecules using docking and molecular dynamics simulations

The role of water molecules in the active site of cytochrome P450 1A2 (CYP1A2) was investigated using an explicit water model to simulate biological environments. Moreover, differences in ligand recognition between the inhibitor α-naphthoflavone (ANF) and the substrate 7-ethoxyresorufin (7ER) in the CYP1A2 complex were examined. More than 200-ns molecular dynamics (MD) simulations were performed for each complex structure of CYP1A2. In the complex structure with 7ER obtained after MD simulation, some water molecules existed in the active site and formed hydrogen bonds between 7ER and some residues. However, in the complex structure with ANF, the hydrogen bond network differed. These results suggest that CYP1A2 requires water molecules in its active site for substrate recognition. The observed differences in the hydrogen bond network in the complex with ANF or 7ER may be due to the fact that ANF is an inhibitor.

Synthesis of Potent and Selective Inhibitors of Aldo-Keto Reductase 1B10 and Their Efficacy against Proliferation, Metastasis, and Cisplatin Resistance of Lung Cancer Cells

Aldo-keto reductase 1B10 (AKR1B10) is overexpressed in several extraintestinal cancers, particularly in non-small-cell lung cancer, where AKR1B10 is a potential diagnostic marker and therapeutic target. Selective AKR1B10 inhibitors are required because compounds should not inhibit the highly related aldose reductase that is involved in monosaccharide and prostaglandin metabolism. Currently, 7-hydroxy-2-(4-methoxyphenylimino)-2H-chromene-3-carboxylic acid benzylamide (HMPC) is known to be the most potent competitive inhibitor of AKR1B10, but it is nonselective. In this study, derivatives of HMPC were synthesized by removing the 4-methoxyphenylimino moiety and replacing the benzylamide with phenylpropylamide. Among them, 4c and 4e showed higher AKR1B10 inhibitory potency (IC50 4.2 and 3.5 nM, respectively) and selectivity than HMPC. The treatments with the two compounds significantly suppressed not only migration, proliferation, and metastasis of lung cancer A549 cells but also metastatic and invasive potentials of cisplatin-resistant A549 cells.