HIV-1整合酶四聚体结构模拟及其活性位点分析

HIV-1复制需要HIV-1整合酶将其环状DNA整合进宿主DNA中,这其中包括2个重要反应,即“3′-加工”和“链转移”,两者均由HIV-1整合酶催化完成.阻断其中的任一反应,都能达到抑制HIV-1复制的目的.因此,了解HIV-1整合酶的完整结构和聚合状态,对深入探讨其作用机理及设计新型抑制剂具有重要的指导作用.然而,迄今为止仅有HIV-1整合酶单独结构域的晶体结构可供参考,而其全酶晶体结构尚未获得解析.本研究利用分子模拟技术,通过蛋白质 蛋白质/DNA分子对接、动力学模拟等方法,构建了全长整合酶四聚体的结构模型、HIV-1 DNA与整合酶复合物的结构模型,进一步从理论上证实HIV-1整合酶是以四聚体形态发挥催化作用,明确“3′-加工”和“链转移”在HIV-1整合酶上的催化位点.同时,通过与作用机理相似的细菌转座子Tn5转座酶等的结构比对,推测HIV-1整合酶的核心结构域中应有第2个Mg^２＋存在,其位置螯合于Asp64与Glu152之间.在HIV-1整合酶结构研究的基础上,有望进一步设计出新的抗艾滋病药物.     

Homology modeling,docking and structure-based virtual screening for new inhibitor identification of Klebsiella pneumoniae heptosyltransferase-III

Abstract

Klebsiella pneumoniae (K. pneumoniae) is a Gram-negative opportunistic pathogen commonly associated with hospital-acquired infections that are often resistant even to antibiotics. Heptosyltransferase (HEP) belongs to the family of glycosyltransferase-B (GT-B) and plays an important in the synthesis of lipopolysaccharides (LPS) essential for the formation of bacterial cell membrane. HEP-III participates in the transfer of heptose sugar to the outer surface of bacteria to synthesize LPS. LPS truncation increases the bacterial sensitivity to hydrophobic antibiotics and detergents, making the HEP as a novel drug target. In the present study, we report the 3D homology model of K. pneumoniae HEP-III and its structure validation. Active site was identified based on similarities with known structures using Dali server, and structure-based pharmacophore model was developed for the active site substrate ADP. The generated pharmacophore model was used as a 3D search query for virtual screening of the ASINEX database. The hit compounds were further filtered based on fit value, molecular docking, docking scores, molecular dynamics (MD) simulations of HEP-III complexed with hit molecules, followed by binding free energy calculations using Molecular Mechanics-Poisson–Boltzmann Surface Area (MM-PBSA). The insights obtained in this work provide the rationale for design of novel inhibitors targeting K. pneumoniae HEP-III and the mechanistic aspects of their binding.

Communicated by Ramaswamy H. Sarma     

New potential inhibitors of mTOR: a computational investigation integrating molecular docking,virtual screening and molecular dynamics simulation

The mTOR (mammalian or mechanistic Target Of Rapamycin), a complex metabolic pathway that involves multiple steps and regulators, is a major human metabolic pathway responsible for cell growth control in response to multiple factors and that is dysregulated in various types of cancer. The classical inhibition of the mTOR pathway is performed by rapamycin and its analogs (rapalogs). Considering that rapamycin binds to an allosteric site and performs a crucial role in the inhibition of the mTOR complex without causing the deleterious side effects common to ATP-competitive inhibitors, we employ ligand-based drug design strategies, such as virtual screening methodology, computational determination of ADME/Tox properties of selected molecules, and molecular dynamics in order to select molecules with the potential to become non-ATP-competitive inhibitors of the mTOR enzymatic complex. Our findings suggest five novel potential mTOR inhibitors, with similar or better properties than the classic inhibitor complex, rapamycin.     

Molecular modeling,dynamics, and an insight into the structural inhibition of cofactor independent phosphoglycerate mutase isoform 1 from Wuchereria bancrofti using cheminformatics and mutational studies

Phosphoglycerate mutase catalyzes the interconversion between 2-phosphoglycerate and 3-phosphoglycerate in the glycolytic and gluconeogenic pathways. They exist in two unrelated forms, that is either cofactor (2,3-diphosphoglycerate) dependent or cofactor-independent. These two enzymes have no similarity in amino acid sequence, tertiary structure, and in catalytic mechanism. Wuchereria bancrofti (WB) contains the cofactor-independent form, whereas other organisms can possess the dependent form or both. Since, independent phosphoglycerate mutase (iPGM) is an essential gene for the survival of nematodes, and it has no sequence or structural similarity to the cofactor-dependent phosphoglycerate mutase found in mammals, it represents an attractive drug target for the filarial nematodes. In this current study, a putative cofactor-iPGM gene was identified in the protein sequence of the WB. In the absence of crystal structure, a three-dimensional structure was determined using the homology modeling approximation, and the most stable protein conformation was identified through the molecular dynamics simulation studies, using GROMACS 4.5. Further, the functional or characteristic residues were identified through the sequence analysis, potential inhibitors were short-listed and validated, and potential inhibitors were ranked using the cheminformatics and molecular dynamics simulations studies, Prime MM-GBSA approach, respectively.     

Stabilization of the Bio-membrane by Small Molecules: Interaction of Trehalose with the Phospholipid Bilayer

Abstract

Anhydrobiotic organisms undergo periods of acute dehydration during their life cycle. It is of interest to understand how the biomembrane remains intact through such stress. A di-saccharide, trehalose, which is metabolised during anhydrobiosis is found to prevent disruption of model membrane systems (1). Molecular modelling techniques are used to investigate the possible mode of interaction of trehalose with a model monolayer. The objective is to maximise hydrogen bonding between the two systems. A phospholipid matrix consisting of l,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC) is chosen to represent the monolayer. The crystal structure of DMPC (2) reveals that there are two distinct conformers designated as A and B. An expansion of the monolayer, coplanar with its surface, results in the trehalose molecule being accomodated in a pocket formed by four B conformers. One glucose ring of the sugar rests on the hydrophobic patch provided by the choline methyls of an A conformer. Five hydrogen bonds are formed involving the phosphate oxygens of three of the surrounding B conformers. The model will be discussed with reference to relevant experimental data on the interaction.     

Molecular dynamics simulation of the docking of substrates to proteins

A simple method is described to perform docking of subtrates to proteins or probes to receptor molecules by a modification of molecular dynamics simulations. The method consists of a separation of the center-of-mass motion of the substrate from its internal and rotational motions, and a separate coupling to different thermal baths for both types of motion of the substrate and for the motion of the receptor. Thus the temperatures and the time constants of coupling to the baths can be arbitrarily varied for these three types of motion, allowing either a frozen or a flexible receptor and allowing control of search rate without disturbance of internal structure. In addition, an extra repulsive term between substrate and protein was applied to smooth the interaction. The method was applied to a model substrate docking onto a model surface, and to the docking of phosphocholine onto immunoglobulin McPC603, in both cases with a frozen receptor. Using transrational temperatures of the substrate in the range of 1300–1700 K and room temperature for the internal degrees of freedom of the substrate, an efficient nontrapping exploratory search (“helicopter view”) is obtained, which visits the correct binding sites. Low energy conformations can then be further investigated by separate search or by dynamic simulated annealing. In both cases the correct minima were identified. The possibility to work with flexible receptors is discussed. © 1994 Wiley-Liss, Inc.     

Discover potential inhibitors for PFKFB3 using 3D-QSAR,virtual screening,molecular docking and molecular dynamics simulation

Abstract

The 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (PFKFB3) is a master regulator of glycolysis in cancer cells by synthesizing fructose-2,6-bisphosphate (F-2,6-BP), a potent allosteric activator of phosphofructokinase-1 (PFK-1), which is a rate-limiting enzyme of glycolysis. PFKFB3 is an attractive target for cancer treatment. It is valuable to discover promising inhibitors by using 3D-QSAR pharmacophore modeling, virtual screening, molecular docking and molecular dynamics simulation. Twenty molecules with known activity were used to build 3D-QSAR pharmacophore models. The best pharmacophore model was ADHR called Hypo1, which had the highest correlation value of 0.98 and the lowest RMSD of 0.82. Then, the Hypo1 was validated by cost value method, test set method and decoy set validation method. Next, the Hypo1 combined with Lipinski's rule of five and ADMET properties were employed to screen databases including Asinex and Specs, total of 1,048,159 molecules. The hits retrieved from screening were docked into protein by different procedures including HTVS, SP and XP. Finally, nine molecules were picked out as potential PFKFB3 inhibitors. The stability of PFKFB3-lead complexes was verified by 40?ns molecular dynamics simulation. The binding free energy and the energy contribution of per residue to the binding energy were calculated by MM-PBSA based on molecular dynamics simulation.     

HIV-1整合酶核心区野生型和F185K突变型活性和溶解性的比较及分子动力学模拟分析

表达纯化了野生型(WT)及F185K突变型HIV-1整合酶核心区蛋白(IN_C),并对二者的溶解性和活性进行了比较．实验结果表明：F185K 突变后IN_C溶解性显著提高,活性有一定程度降低．对WT和F185K IN_C体系进行了1800 ps的分子动力学模拟．模拟结果表明：F185K IN_C功能loop区柔性和蛋白质整体运动性降低,使蛋白质活性降低,F185K突变后盐桥网络的变化驱动了IN_C局部构象改变,引起IN_C表面的部分疏水残基被包埋,亲水残基暴露,相对亲水溶剂可接近面积增大,同时,突变后IN_C与水之间形成氢键的数量增加,与水之间作用加强,以上变化使IN_C溶解性提高．分子动力学模拟与实验结果相吻合．为理解蛋白质溶解性和对蛋白质进行可溶性改造提供了一定的理论依据．     

Homology modeling of human histone deacetylase 10 and design of potential selective inhibitors

Abstract

Histone deacetylases (HDACs) are implicated in the pathology of various cancers, and their pharmacological blockade has proven to be promising in reversing the malignant phenotypes. However, lack of crystal structures of some of the human HDAC isoforms (e.g., HDAC10) hinders the design of the isoform-selective inhibitor. Here, the recently solved X-ray crystal structure of Danio rerio (zebrafish) HDAC10 (Protein Data Bank (PDB) ID; 5TD7, released on 24 May 2017) was retrieved from the PDB and used as a template structure to model the three-dimensional structure of human HDAC10. The overall quality of the best model (M0017) was assessed by computing its z-score—a measure of the deviation of the total energy of the structure with respect to an energy distribution derived from random conformations and by docking of known HDAC10 inhibitors to its catalytic cavity. Furthermore, to identify potential HDAC10-selective inhibitor ligand-based virtual screening was carried out against the ZINC database. The free modeled structure of HDAC10 and its complexes with quisinostat and the highest-ranked compound ZINC19749069 were submitted to molecular dynamics simulation. The comparative analysis of root-mean-squared deviation, root-mean-squared fluctuation, radius of gyration (Rg), and potential energy of these systems showed that HDAC10-ZINC19749069 complex remained the most stable over time. Thus, M0017 could be potentially used for structure-based inhibitor against HDAC10, and ZINC19749069 may provide a scaffold for further optimization.

Communicated by Ramaswamy H. Sarma     

Molecular dynamics simulation and molecular docking studies of 1,4-Dihydropyridines as P-glycoprotein’s allosteric inhibitors

P-glycoprotein (P-gp) is a main factor contributing to multidrug resistance. The effect of this transporter protein on limiting the effectiveness of chemotherapy has been shown by various studies. In a previous report, we synthesized some 14-dihydropyridine (DHP) derivatives as inhibitors of human P-gp. In the present study, a computational approach has been exploited to reveal the main interactions between DHPs and P-gp. In order to do this, homology modeling was performed to obtain a model of the protein. Then, molecular dynamics simulation was used to refine the constructed model of P-gp in the presence of the lipids bilayer. Model validation was performed with several tools. Finally, molecular docking followed by MD simulation of ligand–protein complex was employed to elucidate the binding mode and the dynamical changes of protein with/without DHPs bound. The results emphasized that interaction of the residues Gln912, Ser909, Arg905, Ser474, Val472 with DHPs play a crucial role in the inhibitory of these ligands and this was in a relatively good accordance with the results reported in the experimental studies.     

Molecular dynamics simulation,binding free energy calculation and molecular docking of human D-amino acid oxidase (DAAO) with its inhibitors

Schizophrenia is a mental illness; most affected people live in developing countries, and neither appropriate treatment nor commercial drugs are currently available. One possibility is to inhibit human-d-amino acid oxidase (h-DAAO). In this study, molecular dynamic simulations of the monomer, dimer and tetramer forms of h-DAAO complexed with the inhibitor 3-hydroxyquinolin-2(1H)-one(2) were performed. Seven residues, Leu51, Gln53, Leu215, Tyr228, Ile230, Arg283 and Gly313, were identified as essential for interacting with the inhibitor. Molecular docking of h-DAAO with pyrrole, quinoline and kojic acid derivatives, representing 69 known or potential h-DAAO inhibitors, was also performed. The results indicated that the activity of the inhibitor can be improved by modifying the compounds to have a substituent group capable of interacting with the side chain of Tyr228. Van der Waals interactions of the inhibitor with the hydrophobic pocket of h-DAAO and electrostatic interactions or H-bonds with Arg283 and Gly313 were important elements in determining the efficiency of the inhibitor. These results provide information on the interaction between h-DAAO and its inhibitors at the molecular level and can aid in the design of novel inhibitors against h-DAAO for new drug development in the treatment of schizophrenia.     

Novel virtual lead identification in the discovery of hematopoietic cell kinase (HCK) inhibitors: application of 3D QSAR and molecular dynamics simulation

High level of hematopoietic cell kinase (Hck) is associated with drug resistance in chronic myeloid leukemia. Additionally, Hck activity has also been connected with the pathogenesis of HIV-1 and chronic obstructive pulmonary disease. In this study, three-dimensional (3D) QSAR pharmacophore models were generated for Hck based on experimentally known inhibitors. A best pharmacophore model, Hypo1, was developed with high correlation coefficient (0.975), Low RMS deviation (0.60) and large cost difference (49.31), containing three ring aromatic and one hydrophobic aliphatic feature. It was further validated by the test set (r?=?0.96) and Fisher’s randomization method (95%). Hypo 1 was used as a 3D query for screening the chemical databases, and the hits were further screened by applying Lipinski’s rule of five and ADMET properties. Selected hit compounds were subjected to molecular docking to identify binding conformations in the active site. Finally, the appropriate binding modes of final hit compounds were revealed by molecular dynamics (MD) simulations and free energy calculation studies. Hence, we propose the final three hit compounds as virtual candidates for Hck inhibitors.     

虚拟筛选与新药发现

虚拟筛选是创新药物研究的新方法和新技术,近年来引起了研究机构和制药公司的高度重视,并且已经成为一种与高通量筛选互补的实用化工具,加入到了创新药物研究的工作流程(pipeline)中。本文介绍国际上虚拟筛选及其在创新药物发现中应用的研究进展,特别介绍了我国这方面研究的状况。