Recognition of cyclooxygenase-2 (COX-2) active site by NSAIDs: a computer modelling study.

The energetics and models of COX-2 complexed with nonsteroidal anti-inflammatory drugs (NSAIDs) having different degrees of selectivity for two isoforms of COX (COX-2 and COX-1) have been studied using computer modelling approach. The models are obtained for complexes of NS398 (NS), a selective COX-2 inhibitor; indoprofen (Ind), a non-selective inhibitor; di-tert-butylbenzofurans (DHDMBFs) with substituents at the 5th position: CONH(CH2)2OMe (BF1), CONH-c-Pr (BF2), 3-methylene-gamma-butyrolactonyl (BF3) and oxicams namely, meloxicam (Mel), piroxicam (Pir) and tenoxicam (Ten). These were optimized using molecular mechanics (MM) and molecular dynamics (MD) techniques. The binding energies and structures were compared with pharmacological parameters and available results with COX-1. In case of NS a larger difference in the binding energies between COX-2 and COX-1 was noticed as compared to that of Ind. It also had stronger interaction with His90 and Tyr355 which is considered important for COX-2 selectivity. There was a difference in the compactness at the channel entrance between COX-2 selective and non-selective ligands. Models with DHDMBFs and oxicams showed a similar correlation. The results were used to design a peptide inhibitor, Tyr-Arg-Cys-Ala-delta Phe-Cys (Pept) which could fit better in the COX-2 cavity. As per our MD simulation results this peptide inhibitor showed both higher activity and COX-2 selectivity.

Enzyme selectivity of new cyclooxygenase-2/5 lipoxygenase inhibitors using molecular modeling approach.

We have studied the conformational flexibility of three 5-keto-substituted 7-tert-butyl-2,3-dihydro-3,3-dimethylbenzofurans (DHDMBFs) which show dual cyclooxygenase (COX) and 5-lipoxygenase (LOX) inhibition and are potential candidates as antiinflammatory agents and analgesics. The conformations were studied by systematic search, molecular mechanics (MM) and simulated annealing molecular dynamics (SAMD) techniques. We also studied several structure based parameters and distribution of molecular electrostatic potential (MEP) around these molecules. All the three compounds were docked in the active cavity of cyclooxygenase-2 (COX-2) using graphical and energy grid search techniques. The complex geometries were optimized by MM. The results on conformational flexibility, inter-atomic distances and angles, MEP distribution and points of contacts with peptide side chains in active cavity have been used to understand the mechanistic cause of differential action of these molecules.

Theoretical study of conformational flexibility of tuftsin in vacuum and in aqueous environment.

Conformational flexibility of tuftsin molecule is studied using all-atom based atom-atom potential and systematic search, simulated annealing molecular dynamics (SAMD) and molecular dynamics (MD) techniques. Latter was carried out for 650 pico seconds (ps) using AMBER 4.0 with explicit water in TIP3P model. Number of inter-atomic distances and torsional angles were monitored during SAMD and MD simulation. We found that tuftsin molecule, irrespective of any starting conformation, assumes highly folded structure with strong electrostatic interaction between Lys-2 NH3 and Arg-4 carboxylic group and weak hydrogen bond between Lys-2 CO and Arg-4 NH atoms. It had distorted but stable conformation close to inverse gamma turn.

Conformational flexibility of voltage gated dihydropyridine sensitive calcium channel in hydrated DMPC bilayer.

We have built a model for Ca2+ channel using amino acid sequence from S3 helix of the fourth internal repeat of alpha 1 subunit of dihydropyridine sensitive calcium channel from rabbit skeletal muscle, on the basis of X-ray crystallographic data on four helix bundle. The assembling of the geometry of the pore was achieved using a sixteen residues peptide fragment from short SSI/II loop (residues 1010-1025) which had F1013 and E1014 residues, considered to be important for the drug induced activity of the channel. This had hairpin bend between F1013 to W1016. The drug 2,6-dimethyl 3,5-dicarbomethoxy-4 (2-nitrobenzyl) 1,4 dihydropyridine (DHP) (nifedipine), which is a calcium channel inhibitor used in the treatment of cardiovascular diseases, was introduced, interacting with these two residues via Ca2+ ion. Two more Ca2+ ions were introduced in the pore. The model was incorporated in the bilayer of 36 dimyristoyl phosphatidyl choline (DMPC) molecules with 1201 water molecules and simulated for 200 picoseconds (ps) after equilibration for 120 ps. We also simulated the channel model in vacuum and in aqueous environment for comparison. The latter was unstable after 120 ps. The geometric parameters of the pore are analysed by MOLMOL, PCURVE 3.1 and a special program ANHELIX developed by us. Stability of the pore dimensions during simulations is discussed in this paper.

500 picosecond molecular dynamics simulation of amphiphilic polypeptide Ac(LKKL)4 NHEt with 1,2 di-mysristoyl-sn-glycero-3-phosphorylcholine (DMPC) molecules.

Molecular dynamics (MD) simulation of the interaction between amphiphilic polypeptide Ac(LKKL)4NHEt and 4 DMPC (1,2 di-mysristoyl-sn-glycero-3-phosphorylcholine) molecules has been carried out at 310 K for 500 picoseconds (ps) using AMBER 4.0. Interaction energy and a number of conformational parameters are calculated for the subaveraged coordinates, using P-CURVES 3.1 and our MD trajectory analysis program ANALMD. No significant change in DMPC headgroup conformation was observed. However, the mobility of P atoms was found to be restricted. The chains were quite flexible and their flexibility increased towards the ends. They interacted amongst themselves. The polypeptide remained predominantly in alpha-helical conformation. Leu1 and Lys2 at the N terminus and Leu13 to Leu16 at C terminus assumed non helical conformation and were quite flexible. Average interaction energy between the polypeptide and DMPC molecules was found to be -151.828 kcal*mol-1. The main contributory factor was electrostatic interaction of Lys NH3+ groups with the DMPC phosphates. On an average one Lys chain interacted with 1.5 DMPC molecules. Central region of the polypeptide had better contact with DMPC molecules. A model for the fusogenic properties of the polypeptide is presented on the basis of MD results.

Conformation of nifedipine in hydrated 1,2-di-myristoyl-sn-glycero-3-phosphorylcholine bilayer molecular dynamics simulation.

The conformation of nifedipine, a cardiac and smooth muscle calcium ion channel antagonist is studied in a hydrated bilayer of forty nine l,2-di-myristoyl-sn-glycero-3-phosphorylcholine (DMPC) molecules using molecular dynamics (MD) simulation technique. The simulation was carried out in conditions of constant number, volume and temperature (NVT) at 310 K, which is above the liquid crystalline (Lα) transition temperature of DMPC. The periodic boundary conditions were applied in three-dimensions. Thus the model represented an infinite bilayer. The important geometric parameters characteristic to DMPC and nifedipine molecules were calculated and compared with other theoretical and experimental results pertaining to nifedipine and other related dihydrophyridine (DHP) analogues. Our results suggest that conformational parameters required for antagonist activity are fairly conserved during the interaction of nifedipine with DMPC bilary and bilayer stabilizes the drug conformation in the bioactive form.

Molecular dynamics simulation of hydrated phospholipid bilayers.

Understanding of microscopic behaviour of biological membrane is crucial for designing of molecules to control transport properties of the membranes. Phospholipid-water forms a good model system to study ligand induced structural and dynamical changes in membrane. The review has its main focus on molecular dynamics (MD) simulation of phospholipid bilayers. A brief summary of the current status of structure of phospholipid membranes based on different physico-chemical measurements is given. We discuss here mainly results of MD simulations in the recent years on hydrated phospholipid bilayers and their interaction with ligands. Simulation parameters as: choice of initial system, force fields, protocols for simulation are compared. Main results on: order parameter, head group and chain conformation, water penetration profile, chain tilts, pair-correlation function between atoms of lipid and water, diffusion of ions and ligands are discussed. The review gives application and limitation of MD method for studying lipid water system.

Molecular dynamics study of interaction of dimyristoyl phosphotidyl choline with water.

We present here results of molecular dynamics (MD) simulations on hydrated bilayers of 40 molecules of 1-2-dimyristoyl-sn-glycero-3-phosphatidyl choline (DMPC) in liquid crystalline (Lα) phase using two different models (i) with same (A) conformation for all DMPC molecules, (ii) with alternate rows having different (A and B reported in crystallographic studies on DMPC) conformations. The bilayers were hydrated using 776 and 1064 water molecules. Simulations have been carried out at 310K with AMBER 4·0 program, using united atom force field for 200 pico seconds (ps) after equilibration. During heating and equilibration constant pressure temperature (PT) conditions were maintained while in simulation of equillibrated bilayers constant volume temperature (VT) conditions were used. Subaveraged atomic coordinates were used to calculate geometric parameters of lipid molecules and lipid water interaction. Our results show larger flexibility of polar head group and glycerol region in Lα phase compared to gel or non-hydrated bilayers. Chain disorder was more towards end. Sn-2 chains were more disordered. Use of two types of starting conformations increased disorder. Trans fraction of chain torsional angle was higher in non-hydrated bilayer. However it was more disordered due to ‘swing’ movement of chains because of distortion in torsional angles α2 and 03 due to absence of water molecules. Trans fraction of the chains, order parameter and water penetration showed general agreement with the available experimental results. On the whole MD technique was found to be quite useful for depicting microscopic behaviour of liquid crystalline system and correlating the same with macroscopic changes observed experimentally.

200 picosecond molecular dynamics simulation of interaction of nifedipine with 1-2 dimyristoyl phosphatidylcholine membrane.

Interaction of calcium channel antagonist nifedipine (Nif) with 1-2 dimyristoyl phosphatidylcholine (DMPC) membrane has been studied using molecular dynamics approach. The simulations for one molecule of nifedipine with four DMPC molecules were carried out for 200 pico seconds (ps) using AMBER (Assisted Model Building with Energy Refinement) 3.0 adopted to CYBER 180/930 computer and changes in the structural parameters of DMPC were compared with those for DMPC monolayer (a matrix of nine molecules) optimized separately. Dynamics simulations for the latter had been carried for 40 ps. Our results show that the drug molecule (Nif) penetrates a discrete depth within the phospholipid matrix causing hydrocarbon chains of lipid molecules to swing that makes enough room for the receptor adjacent to the drug molecule.

Molecular mechanics simulation of the interaction of estrogen receptor with estrogen regulatory element.

A model for the interaction of 31 amino acid fragment (protein) from DNA binding domain of human estrogen receptor (hER) with a five base pair DNA sequence 5'GGTCA 3' from estrogen regulatory element (ERE) has been obtained using a step-wise procedure based on structural data on model peptides, DNA binding domain of hER, steric constrains imposed by tetrahedral coordination of the Cys sulphurs with zinc ion and classical secondary structural elements. Structure of the protein as well as its complex with DNA is obtained by energy minimization followed by refinement by molecular mechanics. The complex is stabilized by H-bonds between Lys22, Lys26 and Arg27 with DNA bases G2, T3 and T6. Lys22 also made H-bond with the backbone of G2. The backbone of Cys18 H-bonded with N7 of G1. DNA was in distorted B form and showed evidence of protein-induced conformational changes.

Computer aided study of the interaction of thyroid hormone receptor with DNA.

Interaction of the first zinc finger from human thyroid hormone receptor (finger) with hexanucleotide duplex d(AGGTCA).d(TGACCT) from thyroid regulatory element has been studied by molecular mechanics simulation technique. The structure of the finger as well as its complex with DNA is optimized using constrain of tetrahedral coordination of the zinc ion to Cys sulphurs. The finger is stabilized by series of H--bonds (5 in backbone, 2 in side chains and 2 between backbone and side chains). The complex is stabilized by H-bonds between side chains of Tyr 11, His 12, Tyr 13 and Arg 14 with G2, G3 and G8. DNA is in B form. H--bonding network within DNA is preserved. Opposite strand P-P and Cl'-Cl' distances are changed slightly. There is a systematic change in the backbone torsional angles and sugar pucker. Also, there is an evidence of protein-induced conformational change in DNA.

Stereochemical aspects of interaction of DNA binding domain of human progesterone receptor with d(AGGTCATGCT)2.

Structures of (i) 66 amino-acid fragment (residues 567-633) from DNA binding domain of human progesterone receptor (hPR), (ii) a ten base pair DNA sequence d(AGGTCATGCT)2 from hormone responsive element (HRE) and (iii) a complex of these two are optimised by computer modelling and molecular mechanics technique using extensive steric constraints from secondary structure predictions, comparison with the structures of known metalloproteins, geometric constraints imposed by tetrahedral coordination with the zinc ion and comparison with structures of DNA binding domains of human glucocorticoid and estrogen receptors (hGR and hER). Structure of the complex was obtained using genetic modification data on steroid receptors and general consensus about protein-DNA interaction. DNA is in distorted B conformation. Sequence dependent as well as protein-induced conformation changes are noticed. There is change in propeller twist, buckle and angle between glycosyl bonds. However, H-bonding network is preserved. The complex is stabilized with eighteen hydrogen-bonds, mainly between peptide side-chains and backbone phosphate. There are five specific H-bonds between basic amino acid side chains, Lys 22, Lys 26 and Arg 27, and DNA bases, A1, G3, G16 and A17. Gly 19, Ser 20 and Val 23 are in close proximity of DNA.

Dynamic fluorescence polarization studies on lipid mobilities in phospholipid vesicles in the presence of calcium mediators.

The influence of Ca2+ mediators (nifedipine, verapamil and prostaglandin F2α) on fluorescence polarization of l-anilino-8-napthalene-sulphonate in dipalmitoyl phosphatidylcholine and dimyristoyl phosphatidylcholine liposomes was studied at various temperatures to understand the dynamic behaviour of membrane lipids. We also studied the effect of change in calcium concentration on the fluorescence polarization of the dye in the liposomes. Our results show increase in polarization (indicative of stiffening of the membrane) in the presence of Ca2+ ions. In the case of dimyristoyl phosphatidylcholine liposomes, all 3 drugs caused decrease in fluorescence polarization (increase in fluidity of the membrane) with or without Ca2+ ions in the medium. Contrary to this, in the case of dipalmitoyl phosphatidylcholine liposomes, the fluidization effect is observed for all the 3 drugs in the absence of Ca2+ ions; in the presence of Ca2+ ions stiffening is observed upon addition of nifedipine and verapamil which are antagonists, and fluidization is observed upon addition of prostaglandin F2α· The role of drug-induced fluidity changes in membranes in therapy planning is discussed in the paper.

Computer simulation of the interaction of a3helix of λ Cro protein with DNA and origin of sequence specific recognition.

The conformation of the a3 helix of Cro protein (residues 27–36) of bacteriophage λ is optimised by the damped least square minimization technique, with the steric constraint that Ca atom positions should match the crystallographic data available to date. On the basis of minimization of total interaction and conformation energy, models for complexes of this peptide sequence with heptanucleotide duplexes from native and altered OR3 operator are obtained in the major groove of Β DNA. Analysis of the energetics for 3 sequences of the DNA show that binding strength is derived mainly from the interaction of side chains of the peptide with DNA. Sequence specificity (maximum difference in binding energy for different DNA sequences) is due to hydrogen bonding interaction. A small amount of sequence specificity is derived from non-bonded interaction also. Stereochemical aspects of peptide DNA interaction and their role in DNA recognition are discussed in this paper.