Structural and dynamical properties of a full-length HIV-1 integrase: molecular dynamics simulations

The structural and dynamical properties of the complete full-length structure of HIV-1 integrase were investigated using Molecular Dynamics approach. Simulations were carried out for the three systems, core domain only (CORE), full-length structure without (FULL) and with a Mg2+ (FULL+ION) in its active site, aimed to investigate the difference in the molecular properties of the full-length models due to their different construction procedures as well as the effects of the two ends, C- and N-terminal, on those properties in the core domain. The full-length structure was prepared from the two experimental structures of two-domain fragment. The following properties were observed to differ significantly from the previous reports: (i) relative topology formed by an angle between the three domains; (ii) the cavity size defined by the catalytic triad, Asp64, Asp116, and Glu152; (iii) distances and solvation of the Mg2+; and (iv) conformation of the catalytic residues. In addition, the presence of the two terminal domains decreases the mobility of the central core domain significantly.     

Computational design of novel fullerene analogues as potential HIV-1 PR inhibitors: Analysis of the binding interactions between fullerene inhibitors and HIV-1 PR residues using 3D QSAR, molecular docking and molecular dynamics simulations

A series of experimentally reported as well as computationally designed monoadducts and bisadducts of [60]fullerene analogues have been used in order to analyze the binding interactions between fullerene based inhibitors and HIV-1 PR employing docking studies. MD simulations of ligand-free and the inhibitor bound HIV-1 PR systems complemented the above studies and provided proper input structure of HIV-1 PR in docking simulations. The obtained results revealed a different orientation of the beta-hairpin flaps at these two systems. In inhibitor bound system, the flaps of the enzyme are pulled in toward the bottom of the active site (the closed form) while, in ligand-free system flaps shifted away from the dual Asp25 catalytic site and this system adopts a semi-open form. The structural analysis of these systems at catalytic and flexible flap regions of the HIV-1 PR through the simulation, assisted in understanding the structural preferences of these regions, as well as, the adopted orientations of fullerene derivatives within the active site of the enzyme. Five different combinations of steroelectronic fields of 3D QSAR/CoMSIA models were obtained from the set of biologically evaluated and computationally designed fullerene derivatives (training set=43, test set=6) in order to predict novel compounds with improved inhibition effect. The best 3D QSAR/CoMSIA model yielded a cross validated r(2) value of 0.739 and a non-cross validated r(2) value of 0.993. The derived model indicated the importance of steric (42.6%), electrostatic (12.7%), H-bond donor (16.7%) and H-bond acceptor (28.0%) contributions. The derived contour plots together with de novo drug design were then used as pilot models for proposing the novel analogues with enhanced binding affinities. Such structures may trigger the interest of medicinal chemists for novel HIV-1 PR inhibitors possessing higher bioactivity.     

HIV-1 integrase catalytic core: molecular dynamics and simulated fluorescence decays.

Two molecular dynamics simulations have been carried out on the HIV-1 integrase catalytic core starting from fully determined crystal structures. During the first one, performed in the absence of divalent cation (6-ns long), the catalytic core took on two main conformations. The conformational transition occurs at approximately 3.4 ns. In contrast, during the second one, in the presence of Mg(2+) (4-ns long), there were no such changes. The molecular dynamics simulations were used to compute the fluorescence intensity decays emitted by the four tryptophan residues considered as the only chromophores. The decay was computed by following, frame by frame, the amount of chromophores that remained excited at a certain time after light absorption. The simulation took into account the quenching through electron transfer to the peptide bond and the fluorescence resonance energy transfer between the chromophores. The fit to the experimental intensity decays obtained at 5 degrees C and at 30 degrees C is very good. The fluorescence anisotropy decays were also simulated. Interestingly, the fit to the experimental anisotropy decay was excellent at 5 degrees C and rather poor at 30 degrees C. Various hypotheses such as dimerization and abnormal increase of uncorrelated internal motions are discussed.     

3D QSAR CoMFA/CoMSIA, molecular docking and molecular dynamics studies of fullerene-based HIV-1 PR inhibitors

For the first time, a set of experimentally reported [60] fullerene derivatives were subjected to the 3D-QSAR/CoMFA and CoMSIA studies. The aim of this study is to propose a series of novel [60] fullerene-based inhibitors with optimal binding affinity for the HIV-1 PR enzyme. The position of the template molecule at the cavity of HIV-1 PR was optimized and 3D QSAR models were developed. Relative contributions of steric/electrostatic fields of the 3D-QSAR/CoMFA and CoMSIA models have shown that steric effects govern the bioactivity of the compounds, but electrostatic interactions play also an important role.The de novo drug design Leapfrog simulations provided a series of novel compounds with predicted improved inhibition effect.     

The design of potent HIV-1 integrase inhibitors by a combined approach of structure-based virtual screening and molecular dynamics simulation

Communicated by Ramaswamy H. Sarma     

Structure and internal dynamics of the bovine pancreatic trypsin inhibitor in aqueous solution from long-time molecular dynamics simulations

Structural and dynamic properties of bovine pancreatic trypsin inhibitor (BPTI) in aqueous solution are investigated using two molecular dynamics (MD) simulations: one of 1.4 ns length and one of 0.8 ns length in which atom-atom distance bounds derived from NMR spectroscopy are included in the potential energy function to make the trajectory satisfy these experimental data more closely. The simulated properties of BPTI are compared with crystal and solution structures of BPTI, and found to be in agreement with the available experimental data. The best agreement with experiment was obtained when atom-atom distance restraints were applied in a time-averaged manner in the simulation. The polypeptide segments found to be most flexible in the MD simulations coincide closely with those showing differences between the crystal and solution structures of BPTI. © 1995 Wiley-Liss, Inc.     

Toward novel HIV-1 integrase binding inhibitors: molecular modeling, synthesis, and biological studies

The identification of a novel hit compound as integrase binding inhibitor has been accomplished by means of virtual screening techniques. A small family of structurally related molecules has been synthesized and biologically evaluated with one of the compounds showing an IC(50)=12 microM.     

Ab initio molecular dynamics simulations of beta-D-glucose and beta-D-xylose degradation mechanisms in acidic aqueous solution

Ab initio molecular dynamics simulations were employed to investigate, with explicit solvent water molecules, beta-D-glucose and beta-D-xylose degradation mechanisms in acidic media. The rate-limiting step in sugar degradation was found to be protonation of the hydroxyl groups on the sugar ring. We found that the structure of water molecules plays a significant role in the acidic sugar degradation pathways. Firstly, a water molecule competes with the hydroxyl group on the sugar ring for protons. Secondly, water forms hydrogen bonds with the hydroxyl groups on the sugar rings, thus weakening the C-C and C-O bonds (each to a different degree). Note that the reaction pathways could be altered due to the change of relative stability of the C-C and C-O bonds. Thirdly, water molecules that are hydrogen-bonded to sugar hydroxyls could easily extract a proton from the reaction intermediate, terminating the reaction. Indeed, the sugar degradation pathway is complex due to multiple protonation probabilities and the surrounding water structure. Our experimental data support multiple sugar acidic degradation pathways.     

Design and synthesis of specific inhibitors of the 3'-processing step of HIV-1 integrase

The novel dinucleotide 5'-phosphate, [(L,D)-pIsodApdC], discovered in our laboratory, is a strong inhibitor of HIV-1 integrase for both the 3'-processing and the strand transfer steps. The rationale used in this molecular design was that residues immediately upstream of the dinucleotide cleavage site in the 3'-processing step might provide critical recognition/binding sites on integrase. The rationale for the second type of inhibitors was based on the elimination products (linear and cyclic dinucleotides) of 3'-processing. However, while the linear dinucleotide 5'-phosphate (pdGpdT) was active, its cyclic counterpart was inactive against both wild-type and mutant HIV integrase.     

Improved prediction of HIV-1 protease-inhibitor binding energies by molecular dynamics simulations

Background  

The accurate prediction of enzyme-substrate interaction energies is one of the major challenges in computational biology. This study describes the improvement of protein-ligand binding energy prediction by incorporating protein flexibility through the use of molecular dynamics (MD) simulations.     

Self-association behaviour of atactic polymethacrylic acid in aqueous solution investigated by atomistic molecular dynamics simulations

The self-association behaviour of atactic poly(methacrylic acid) (a-PMA) in water was investigated by atomistic molecular dynamics (MD) simulations. Simulations show that interchain association of a-PMA occurs only in its un-neutralised form, by hydrogen bonding between –COOH groups, which is in agreement with the experimental observation. Chain conformations, dihedral angle distributions, hydration behaviour, scattering structure factor and enthalpy-of-hydration (i.e. aqueous solvation) were analysed as a function of concentration for un-neutralised PMA, across dilute to concentrated regimes. The average 〈R_g〉 of the chain remains unaffected in solution and also for amorphous undissolved a-PMA phase, confirming the occurrence of the approximate theta-solution condition for the first time, as revealed by simulations, in a polar hydrogen-bonding polymer aqueous solution. Chain hydration behaviour and scattering structure factor show significant changes in concentrated regime. Scattering intensity collapse occurs in concentrated PMA solution, due to the existence of the swollen regime captured for the first time by explicit-MD-simulations. The hydration of PMA is driven by H-bonding, specifically between H atoms of the COOH groups and O atoms of water molecules in the closest coordination shell. The enthalpy of hydration of PMA is dominated by PMA–water interactions (charges and H-bonding). The thermodynamic contributions of PMA–PMA and PMA–water interactions towards the electrostatics as well as the dispersion components of the total solvation-enthalpy become more favourable than water–water interactions.     

De novo design and synthesis of HIV-1 integrase inhibitors

Existing AIDS therapies are out of reach for most HIV-infected people in developing countries and, where available, they are limited by their toxicity and their cost. New anti-HIV agents are needed urgently to combat emerging viral resistance and reduce the side effects associated with currently available drugs. Toward this end, LeapFrog, a de novo drug design program was used to design novel, potent, and selective inhibitors of HIV-1 integrase. The designed compounds were synthesized and tested for in vitro inhibition of HIV-1 integrase. Out of the 25 compounds that were designed, and synthesized, four molecules (compounds 23, 26, 43, and 59) showed moderate to low inhibition of HIV-1 integrase for 3'-processing and 3'-strand transfer activities. Nonetheless, these compounds possess structural features not seen in known HIV-1 integrase inhibitors and thus can serve as excellent leads for further optimization of anti-HIV-1 integrase activity.     

Design and synthesis of novel dihydroquinoline-3-carboxylic acids as HIV-1 integrase inhibitors

Previously, we discovered linomide analogues as novel HIV-1 integrase (IN) inhibitors. Here, to make possible structure–activity relationships, we report on the design and synthesis of a series of substituted dihydroquinoline-3-carboxylic acids. The crystal structure of the representative compound 2c has also been solved. Among the eight new analogues, 2e showed a potency in inhibiting IN strand transfer catalytic activity similar to the reference diketo acid inhibitor L-731,988 (IC₅₀ = 0.9 μM vs. 0.54 μM, for 2e and L-731,988, respectively). Furthermore, none of the compounds showed significant cytotoxicity in two tested cancer cell lines. These compounds represent an interesting prototype of IN inhibitors, potentially involved in a metal chelating mechanism, and further optimization is warranted.     

Full length Vpu from HIV-1: combining molecular dynamics simulations with NMR spectroscopy

Based on structures made available by solution NMR, molecular models of the protein Vpu from HIV-1 were built and refined by 6 ns MD simulations in a fully hydrated lipid bilayer. Vpu is an 81 amino acid type I integral membrane protein encoded by the human immunodeficiency virus type-1 (HIV-1) and closely related simian immunodeficiency viruses (SIVs). Its role is to amplify viral release. Upon phosphorylation, the cytoplasmic domain adopts a more compact shape with helices 2 and 3 becoming almost parallel to each other. A loss of helicity for several residues belonging to the helices adjacent to both ends of the loop region containing serines 53 and 57 is observed. A fourth helix, present in one of the NMR-based structures of the cytoplasmic domain and located near the C-terminus, is lost upon phosphorylation.     

The dynamics of interconverting D- and E-forms of the HIV-1 integrase N-terminal domain

The N-terminal domain (NTD) of HIV-1 integrase adopts two inter-converting forms (D- and E-) due to their specific coordination of a Zn²⁺ ion by an HHCC motif. Mutational studies on NTD have suggested the importance of conformational transition in regulating the functions of tetramers and dimers of HIV-1 integrase. This study explores the stability and dynamics of native NTD forms and the conformational transition between D- and E-forms using molecular dynamics simulations elucidating their role in regulation of viral and host DNA integration. Simulation of native forms of NTD revealed stable dynamics. Transition studies between D- and E-forms using conventional molecular dynamics simulations for 50 ns partially revealed conformational change towards the target during D- to -E simulation (the extension of α1-helix), which failed in the E- to -D simulation. This could be attributed to the existence of the D-form (?1,945.907 kCal/mol) in higher energy than the E-form (?2,002.383 kCal/mol). The conformational transition pathway between these two states was explored using targeted molecular dynamics simulations. Analysis of the targeted molecular dynamics trajectories revealed conformations closer to the experimentally-reported intermediate form of an NTD during the transition phase. The role of Met22 in stabilizing the E-form was studied by simulating the E-form with Met22Ala mutation, revealing a highly dynamic α1-helix as compared to the native form. The present study reveals the significant role of the Zn²⁺ ion-coordinated HHCC motif and its interaction with Met22 as the basis for understanding the biological implications of D- and E-forms of the NTD in regulating integration reaction.     

Peptide inhibitors of HIV-1 integrase dissociate the enzyme oligomers.

Integration of HIV-1 genome into host cell chromosome is mediated by viral integrase (IN). The IN catalytic core (CC, IN(50-212)) dimerizes through mutual interactions of its alpha1 and alpha5 helices. Peptides INH1 and INH5 reproducing these helix sequences strongly inhibited IN. For instance, an IC(50) of 80 nM was determined for INH5 in integration assays using wild-type IN (wtIN). In size exclusion chromatography, INH1 and INH5 perturbed the association-dissociation equilibrium of both dmIN (IN(1-288)/F185K/C280S) and CC, leading to monomers as surviving species, while in circular dichroism, binding of peptides to dmIN altered the protein conformation. Thus, enzyme deactivation, subunit dissociation, and protein unfolding are events which parallel one another. The target of INH5 in the enzyme was then identified. In fluorescence spectroscopy, C(0.5) values of 168 and 44 nM were determined for the binding affinity of INH5 to IN and CC, respectively, at 115 nM subunit concentration, while interaction of INH5 with INH1 was found stronger than interaction of INH5 with itself (23 times larger in term of dissociation constants). These results strongly suggested that the alpha1 helix is the privileged target of INH5. The latter could serve as a lead for the development of new chemotherapeutic agents against HIV-1.     

Discovery of small-molecule HIV-1 fusion and integrase inhibitors oleuropein and hydroxytyrosol: part II. integrase inhibition

We report molecular modeling and functional confirmation of Ole and HT binding to HIV-1 integrase. Docking simulations identified two binding regions for Ole within the integrase active site. Region I encompasses the conserved D64-D116-E152 motif, while region II involves the flexible loop region formed by amino acid residues 140-149. HT, on the other hand, binds to region II. Both Ole and HT exhibit favorable interactions with important amino acid residues through strong H-bonding and van der Waals contacts, predicting integrase inhibition. To test and confirm modeling predictions, we examined the effect of Ole and HT on HIV-1 integrase activities including 3'-processing, strand transfer, and disintegration. Ole and HT exhibit dose-dependent inhibition on all three activities, with EC(50)s in the nanomolar range. These studies demonstrate that molecular modeling of target-ligand interaction coupled with structural-activity analysis should facilitate the design and identification of innovative integrase inhibitors and other therapeutics.     

Molecular dynamics simulations of the two disaccharides of hyaluronan in aqueous solution

Hyaluronan is an unusually stiff polymer when in aqueous solution,which has important consequences for its biological function.Molecular dynamics simulations of hyaluronan disaccharides havebeen performed, with explicit inclusion of water, to determinethe molecular basis of this stiffness, and to investigate thedynamics of the glycosidic linkages. Our simulations revealthat stable sets of hydrogen bonds frequently connect the neighboringresidues of hyaluronan. Water caging around the glycosidic linkagewas observed to increase the connectivity between sugars, andfurther constrain them. This, we propose, explains the unusualstiffness of polymeric hyaluronan. It would allow the polysaccharideto maintain local secondary structure, and occupy large solutiondomains consistent with the visco-elastic nature of hyaluronan.Simulations in water showed no significant changes on inclusionof the exo-anomeric effect. This, we deduced, was due to hyaluronandisaccharides ordering first shell water molecules. In somecases these waters were observed to transiently induce con-formationalchange, by breaking intramolecular hydrogen bonds. conformation hyaluronan hydrogen bonds molecular dynamics water     

Characterization of molecular structures and properties of polyurethanes using molecular dynamics simulations

Thermotropic polyurethanes with mesogenic groups in side chains were prepared from two diisocyanates and four diols with stoichiometric ratios of reactive isocyanate (NCO) and hydroxy (OH) groups. Their thermal behavior was determined by differential scanning calorimetry. The effect of structure modifications of the diisocyanates and diols, in particular changes in the mesogen, were investigated. Introduction of mesogenic segments into the polymers suppresses the ordering. Stiff end substituents (phenyl and alkoxy groups) of the mesogens stabilize the mesophases to such an extent that the negative influence of long polymer chains is compensated and the liquid-crystalline properties are recovered. All-atom molecular dynamics simulations in the Cerius² modeling environment were carried out to characterize the structures of the polymers. Analysis of the dynamic trajectories at 20, 100, 120 and 170 °C revealed changes in conformation of macromolecules, which correlate with DSC measurements.Figure Example of structure relaxation of D4/TDI molecule at indicated simulation times (temperature 20 °C): a complete structure; b backbone structure; c top view of molecule     

Conformation of endothelin in aqueous ethylene glycol determined by 1H-NMR and molecular dynamics simulations

The solution conformation of a 21-residue vasoconstrictor peptide endothelin-1 (ET-1) in water-ethylene glycol has been determined by two-dimensional 1H-NMR spectroscopy and constrained molecular dynamics simulations. The N-terminus (residues 1-4) appears to undergo conformational averaging and no single structure consistent with the NMR constraints could be found for this region. Residues 5-8 form a turn, and residues 9-16 exist in a helical conformation. A flexible 'hinge' between residues 8-9 allows various orientations of the turn relative to the helix. Another 'hinge' at residue 17 connects the extended C-terminus to the bicyclic core region (residues 1-15). Residues important for binding and biological activity form a contiguous surface on one side of the helix, with the two disulfides extending from the other side of the helix.