Dynamics of water molecules in the active-site cavity of human cytochromes P450

We have studied the dynamics of water molecules in six crystal structures of four human cytochromes P450, 2A6, 2C8, 2C9, and 3A4, with molecular dynamics simulations. In the crystal structures, only a few water molecules are seen and the reported sizes of the active-site cavity vary a lot. In the simulations, the cavities are completely filled with water molecules, although with approximately 20% lower density than in bulk water. The 2A6 protein differs from the other three in that it has a very small cavity with only two water molecules and no exchange with the surroundings. The other three proteins have quite big cavities, with 41 water molecules on average in 2C8 and 54-58 in 2C9 and 3A4, giving a water volume of 1500-2100 A3. The two crystal structures of 2C9 differ quite appreciably, whereas those of 3A4 are quite similar. The active-site cavity is connected to the surroundings by three to six channels, through which there is a quite frequent exchange of water molecules (one molecule is exchanged every 30-200 ps), except in 2A6. Most of the channels are observed also in the crystal structures, but two to three channels in each protein open only during the simulations. There are no water molecules close to the heme iron ion in these simulations of the high-spin ferric state (the average distance to the closest water molecule is 3.3-5 A), and there are few ordered water molecules in the active sites, none of which is conserved in all proteins.     

Prediction and observation of isostructurality induced by solvent incorporation in multicomponent crystals

The crystal structures of two pharmaceutical molecules-carbamazepine and its 10,11-dihydro derivative-with acetic acid have been successfully predicted by computational methods. While the crystalline structure of the former was known a priori, no structural information was available for the latter. Possible crystal structures were generated in silico before any experimental work was performed. Although the crystal structures of the pure drug molecules are very different, incorporation of acetic acid in their crystal lattices results in isomorphic products.     

On the interplay between theory and experiment in molecular structure problems: Some case studies

Some brief, general comments on the concept of molecular structure will be given. Some important points connected to the use of models in the interpretation of molecular structures will also be mentioned. The main theme in one part of the presentation will be the cooperative efforts made by experimentalists (mainly spectroscopists) and theoreticians (computational chemists) in order to measure, predict, and analyze the perturbations on the cyclopropane ring by different substituents. The aim is to demonstrate that ab initio calculations of a certain quality can constitute an important support for experimental studies in that they are able to discriminate between different models that otherwise are equally probable. The second part of the presentation will be concerned with a class of molecules that gives both experimental structural chemists and computational chemists a great challenge, namely, the metallocenes. A discussion of some of the grave discrepancies between theory and experiment regarding their geometry will be given.     

Analysis of ligand-bound water molecules in high-resolution crystal structures of protein-ligand complexes

We have performed a comprehensive analysis of water molecules at the protein-ligand interfaces observed in 392 high-resolution crystal structures. There are a total of 1829 ligand-bound water molecules in these 392 complexes; 18% are surface water molecules, and 72% are interfacial water molecules. The number of ligand-bound water molecules in each complex structure ranges from 0 to 21 and has an average of 4.6. Of these interfacial water molecules, 76% are considered to be bridging water molecules, characterized by having polar interactions with both ligand and protein atoms. Among a number of factors that may influence the number of ligand-bound water molecules, the polar van der Waals (vdw) surface area of ligands has the highest Pearson linear correlation coefficient of 0.63. Our regression analysis predicted that one more ligand-bound water molecule is expected for every additional 24 A2 in the polar vdw surface area of the ligand. In contrast to the observation that the resolution is the primary factor influencing the number of water molecules in crystallographic models of proteins, we found that there is only a weak relationship between the number of ligand-bound water molecules and the resolution of the crystal structures. An analysis of the isotropic B factors of buried ligand-bound water molecules suggested that, when water molecules have fewer than two polar interactions with the protein-ligand complex, they are more mobile than protein atoms in the crystal structures; when they have more than three polar interactions, they are significantly less mobile than protein atoms.     

Surface protonation at the rutile (110) interface: explicit incorporation of solvation structure within the refined MUSIC model framework

The detailed solvation structure at the (110) surface of rutile (alpha-TiO2) in contact with bulk liquid water has been obtained primarily from experimentally verified classical molecular dynamics (CMD) simulations of the ab initio-optimized surface in contact with SPC/E water. The results are used to explicitly quantify H-bonding interactions, which are then used within the refined MUSIC model framework to predict surface oxygen protonation constants. Quantum mechanical molecular dynamics (QMD) simulations in the presence of freely dissociable water molecules produced H-bond distributions around deprotonated surface oxygens very similar to those obtained by CMD with nondissociable SPC/E water, thereby confirming that the less computationally intensive CMD simulations provide accurate H-bond information. Utilizing this H-bond information within the refined MUSIC model, along with manually adjusted Ti-O surface bond lengths that are nonetheless within 0.05 A of those obtained from static density functional theory (DFT) calculations and measured in X-ray reflectivity experiments (as well as bulk crystal values), give surface protonation constants that result in a calculated zero net proton charge pH value (pHznpc) at 25 degrees C that agrees quantitatively with the experimentally determined value (5.4+/-0.2) for a specific rutile powder dominated by the (110) crystal face. Moreover, the predicted pHznpc values agree to within 0.1 pH unit with those measured at all temperatures between 10 and 250 degrees C. A slightly smaller manual adjustment of the DFT-derived Ti-O surface bond lengths was sufficient to bring the predicted pHznpcvalue of the rutile (110) surface at 25 degrees C into quantitative agreement with the experimental value (4.8+/-0.3) obtained from a polished and annealed rutile (110) single crystal surface in contact with dilute sodium nitrate solutions using second harmonic generation (SHG) intensity measurements as a function of ionic strength. Additionally, the H-bond interactions between protolyzable surface oxygen groups and water were found to be stronger than those between bulk water molecules at all temperatures investigated in our CMD simulations (25, 150 and 250 degrees C). Comparison with the protonation scheme previously determined for the (110) surface of isostructural cassiterite (alpha-SnO2) reveals that the greater extent of H-bonding on the latter surface, and in particular between water and the terminal hydroxyl group (Sn-OH) results in the predicted protonation constant for that group being lower than for the bridged oxygen (Sn-O-Sn), while the reverse is true for the rutile (110) surface. These results demonstrate the importance of H-bond structure in dictating surface protonation behavior, and that explicit use of this solvation structure within the refined MUSIC model framework results in predicted surface protonation constants that are also consistent with a variety of other experimental and computational data.     

Ab Initio prediction of possible crystal structures for general organic molecules

The performance of a new crystal packing procedure for the ab initio prediction of possible molecular crystal structures is presented. The method is based upon only molecular information, i.e., no unit cell parameters are assumed to be known. The search for the global crystal energy minimum and all local minima inside an energy window is derived from Monte Carlo simulated annealing methods and has been applied to various organic molecules containing heteroatoms and polar groups. A systematic evaluation of the search method and of the quality of the potential energy function has been established. It is demonstrated that the packing of general organic molecules is possible even with standard force fields like CHARMM provided that the charges defining the electrostatic interactions are based upon physical models rather than transferable empirical parameters. Concepts of crystal packing that were based till now upon assumptions and speculations could be proved or disproved by solving directly the extended global optimization problem related to crystal packing. Crystal structures of molecules as complex as those treated in this article have not been, till now, predicted by a computational approach. In one case, a disagreement between the predicted and experimental structure was evident and, based upon the computations, we suspect that the published structure is the wrong one. © 1992 by John Wiley & Sons, Inc.     

A new model for simulating 3-d crystal growth and its application to the study of antifreeze proteins

A novel computational technique for modeling crystal formation has been developed that combines three-dimensional (3-D) molecular representation and detailed energetics calculations of molecular mechanics techniques with the less-sophisticated probabilistic approach used by statistical techniques to study systems containing millions of molecules undergoing billions of interactions. Because our model incorporates both the structure of and the interaction energies between participating molecules, it enables the 3-D shape and surface properties of these molecules to directly affect crystal formation. This increase in model complexity has been achieved while simultaneously increasing the number of molecules in simulations by several orders of magnitude over previous statistical models. We have applied this technique to study the inhibitory effects of antifreeze proteins (AFPs) on ice-crystal formation. Modeling involving both fish and insect AFPs has produced results consistent with experimental observations, including the replication of ice-etching patterns, ice-growth inhibition, and specific AFP-induced ice morphologies. Our work suggests that the degree of AFP activity results more from AFP ice-binding orientation than from AFP ice-binding strength. This technique could readily be adapted to study other crystal and crystal inhibitor systems, or to study other noncrystal systems that exhibit regularity in the structuring of their component molecules, such as those associated with the new nanotechnologies.     

Predictability of the polymorphs of small organic compounds: crystal structure predictions of four benchmark blind test molecules

Predicting the crystal structure of an organic molecule from first principles has been a major challenge in physical chemistry. Recently, the application of Density Functional Theory including a dispersive energy correction (the DFT(d) method) has been shown to be a reliable method for predicting experimental structures based purely on their ranking according to lattice energy. Further validation results of the application of the DFT(d) method to four organic molecules are presented here. The compounds were targets (labelled molecule II, VI, VII and XI) in previous blind tests of crystal structure prediction, and their structures proved difficult to predict. However, this study shows that the DFT(d) approach is capable of predicting the solid state structures of these small molecules. For molecule VII, the most stable (rank 1) predicted crystal structure corresponds to the experimentally observed structure. For molecule VI, the rank 1, 2 and 3 predicted structures correspond to the three experimental polymorphs, forms I, III and II, respectively. For molecules II and XI, their rank 1 predicted structures are energetically more stable than those corresponding to the experimental crystal structures, and were not found amongst the structures submitted by the participants in the blind tests. The rank 1 structure of molecule II is predicted to exist under high pressure, whilst the rank 1 structure predicted for molecule XI has the same space group and hydrogen bonding pattern as observed in the crystal of 1-amino-1-methyl-cyclopropane, which is structurally related to molecule XI. The experimental crystal structure of molecule II corresponds to the rank 4 prediction, 0.8 kJ mol(-1) above the global minimum structure, and the experimental structure of molecule XI corresponds to the rank 2 prediction, 0.4 kJ mol(-1) above the global minimum.     

A conformationally restricted guanosine analog reveals the catalytic relevance of three structures of an RNA enzyme

Recent studies indicate that RNA function can be enhanced by the incorporation of conformationally restricted nucleotides. Herein, we use 8-bromoguanosine, a nucleotide analog with an enforced syn conformation, to elucidate the catalytic relevance of ribozyme structures. We chose to study the lead-dependent ribozyme (leadzyme) because structural models derived from NMR, crystal, and computational (MC-Sym) studies differ in which of the three active site guanosines (G7, G9, or G24) have a syn glycosidic torsion angle. Kinetic assays were carried out on 8BrG variants at these three guanosine positions. These data indicate that an 8BrG24 leadzyme is hyperactive, while 8BrG7 and 8BrG9 leadzymes have reduced activity. These findings support the computational model of the leadzyme, rather than the NMR and crystal structures, as being the most relevant to phosphodiester bond cleavage.     

New simulation model of multicomponent crystal growth and inhibition

We review a novel computational model for the study of crystal structures both on their own and in conjunction with inhibitor molecules. The model advances existing Monte Carlo (MC) simulation techniques by extending them from modeling 3D crystal surface patches to modeling entire 3D crystals, and by including the use of "complex" multicomponent molecules within the simulations. These advances makes it possible to incorporate the 3D shape and non-uniform surface properties of inhibitors into simulations, and to study what effect these inhibitor properties have on the growth of whole crystals containing up to tens of millions of molecules. The application of this extended MC model to the study of antifreeze proteins (AFPs) and their effects on ice formation is reported, including the success of the technique in achieving AFP-induced ice-growth inhibition with concurrent changes to ice morphology that mimic experimental results. Simulations of ice-growth inhibition suggest that the degree of inhibition afforded by an AFP is a function of its ice-binding position relative to the underlying anisotropic growth pattern of ice. This extended MC technique is applicable to other crystal and crystal-inhibitor systems, including more complex crystal systems such as clathrates.     

Interpretation of protein/ligand crystal structure using QM/MM calculations: case of HIV-1 protease/metallacarborane complex

Deltahedral metallacarborane compounds have recently been discovered as potent, specific, stable, and nontoxic inhibitors of HIV-1 protease (PR), the major target for AIDS therapy. The 2.15 A-resolution X-ray structure has exhibited a nonsymmetrical binding of the parental compound [Co(3+)-(C2B9H11)2](-) (GB-18) into PR dimer and a symmetrical arrangement in the crystal of two PR dimer complexes into a tetramer. In order to explore structural and energetic details of the inhibitor binding, quantum mechanics coupled with molecular mechanics approach was utilized. Realizing the close positioning of anionic inhibitors in the active site cavity, the possibility of an exchange of structural water molecules Wat50 and Wat128 by Na+ counterions was studied. The energy profiles for the rotation of the GB-18 molecules along their longitudinal axes in complex with PR were calculated. The results show that two Na+ counterions are present in the active site cavity and provide energetically favorable and unfavorable positions for carbon atoms within the carborane cages. Eighty-one rotamer combinations of four molecules of GB-18 bound to PR out of 4 x 10(5) are predicted to be highly populated. These results lay ground for further calculations of interaction energies between GB-18 and amino acids of PR active site and will make it possible to interpret computationally the binding of similar metallacarborane molecules to PR as well as to resistant PR variants. Moreover, this computational tool will allow the design of new, more potent metallacarborane-based HIV-1 protease inhibitors.     

Complexes formed between the quadridentate, heterocyclic molecules 6,6'-bis-(5,6-dialkyl-1,2,4-triazin-3-yl)-2,2'-bipyridine (BTBP) and lanthanides(III): implications for the partitioning of actinides(III) and lanthanides(III)

New hydrophobic, tetradentate nitrogen heterocyclic reagents, 6,6'-bis-(5,6-dialkyl-1,2,4-triazin-3-yl)-2,2'-bipyridines (BTBPs) have been synthesised. These reagents form complexes with lanthanides and crystal structures with 11 different lanthanides have been determined. The majority of the structures show the lanthanide to be 10-coordinate with stoichiometry [Ln(BTBP)(NO3)3] although Yb and Lu are 9-coordinate in complexes with stoichiometry [Ln(BTBP)(NO3)2(H2O)](NO3). In these complexes the BTBP ligands are tetradentate and planar with donor nitrogens mutually cisi.e. in the cis, cis, cis conformation. Crystal structures of two free molecules, namely C2-BTBP and CyMe4-BTBP have also been determined and show different conformations described as cis, trans, cis and trans, trans, trans respectively. A NMR titration between lanthanum nitrate and C5-BTBP showed that two different complexes are to be found in solution, namely [La(C5-BTBP)2]3+ and [La(C5-BTBP)(NO3)3]. The BTBPs dissolved in octanol were able to extract Am(III) and Eu(III) from 1 M nitric acid with large separation factors.     

Tosyl esters of cinchonidine and cinchonine alkaloids: the structure–reactivity relationship in the hydrolysis to 9‐epibases

In the crystal structures of the diastereoisomers of O‐tosylcinchonidine [(9R)‐cinchon‐9‐yl 4‐methylbenzenesulfonate], (I), and O‐tosylcinchonine [(9S)‐cinchon‐9‐yl 4‐methylbenzenesulfonate], (II), both C₂₆H₂₈N₂O₃S, both molecules are in an anti‐closed conformation and, in each case, the position of the aryl ring of the tosylate system is influenced by an intramolecular C—H...O hydrogen bond. The molecular packing in (I) is influenced by weak intermolecular C—H...O and C—H...π interactions. The crystal structure of (II) features C—H...π interactions and van der Waals forces only. The computational investigations using RHF/6–31G** ab initio and AM1 semi‐empirical methods performed for (I) and (II) and their protonated species show that the conformational and energetic parameters of the molecules are correlated with differences in their reactivity in hydrolysis to the corresponding 9‐epibases.     

Noble-Noble Strong Union: Gold at Its Best to Make a Bond with a Noble Gas Atom

This Review presents the current status of the noble gas (Ng)-noble metal chemistry, which began in 1977 with the detection of AuNe⁺ through mass spectroscopy and then grew from 2000 onwards; currently, the field is in a somewhat matured state. On one side, modern quantum chemistry is very effective in providing important insights into the structure, stability, and barrier for the decomposition of Ng compounds and, as a result, a plethora of viable Ng compounds have been predicted. On the other hand. experimental achievement also goes beyond microscopic detection and characterization through spectroscopic techniques and crystal structures at ambient temperature; for example, (AuXe₄)²⁺(Sb₂F₁₁⁻)₂ have also been obtained. The bonding between two noble elements of the periodic table can even reach the covalent limit. The relativistic effect makes gold a very special candidate to form a strong bond with Ng in comparison to copper and silver. Insertion compounds, which are metastable in nature, depending on their kinetic stability, display an even more fascinating bonding situation. The degree of covalency in Ng–M (M=noble metal) bonds of insertion compounds is far larger than that in non-insertion compounds. In fact, in MNgCN (M=Cu, Ag, Au) molecules, the M−Ng and Ng−C bonds might be represented as classical 2c–2e σ bonds. Therefore, noble metals, particularly gold, provide the opportunity for experimental chemists to obtain sufficiently stable complexes with Ng at room temperature in order to characterize them by using experimental techniques and, with the intriguing bonding situation, to explore them with various computational tools from a theoretical perspective. This field is relatively young and, in the coming years, a lot of advancement is expected experimentally as well as theoretically.     

The crystalline structures of carboxylic acid monolayers adsorbed on graphite

X-ray and neutron diffraction have been used to investigate the formation of solid crystalline monolayers of all of the linear carboxylic acids from C(6) to C(14) at submonolayer coverage and from C(8) to C(14) at multilayer coverages, and to characterize their structures. X-rays and neutrons highlight different aspects of the monolayer structures, and their combination is therefore important in structural determination. For all of the acids with an odd number of carbon atoms, the unit cell is rectangular of plane group pgg containing four molecules. The members of the homologous series with an even number of carbon atoms have an oblique unit cell with two molecules per unit cell and plane group p2. This odd-even variation in crystal structure provides an explanation for the odd-even variation observed in monolayer melting points and mixing behavior. In all cases, the molecules are arranged in strongly hydrogen-bonded dimers with their extended axes parallel to the surface and the plane of the carbon skeleton essentially parallel to the graphite surface. The monolayer crystal structures have unit cell dimensions similar to certain close-packed planes of the bulk crystals, but the molecular arrangements are different. There is a 1-3% compression on increasing the coverage over a monolayer.     

系列双核Ln(III)配合物的晶体结构和磁性

合成了分子式为[Ln~2(phen)~2L],phen=C~1~2H~8N~2[ALn=Nd,L=(CH~3COO)~4(ONO~2)~2,BLn=Sm,L=(C~6H~5COO)~6,CLn=Eu,L=(C~6H~5COO)~6]3种同双核配合物。用X射线四圆衍射仪测定了3种化合物的结构。在化合物A分子中,2个Nd(III)原子由4个CH~3COO^-基团桥联,以phen和ONO~2^-为端基,构成了一个具有C~2对称性的双核分子。配合物B和C具有完全相同的结构,它们是以4个苯甲酸根为桥,2个phen和2个C~6H~5COO^-为端基的中心对称双核分子,其中6个苯甲酸根的成键状态可分为3种状况。在3种化合物中,每个Ln均为9配位,呈不规则多面体。Ln-Ln距离,A为0.397nm,B和C均为0.405nm。测定了各配合物的变温磁化率,通过对磁性质研究,发现化合物A在低温下具有反铁磁物质行为,并由理论拟合,求得了磁参数g,J值。     

Structural characterization of an anhydrous polymorph of paclitaxel by solid-state NMR

The three-dimensional structure of a unique polymorph of the anticancer drug paclitaxel (Taxol) is established using solid state NMR (SSNMR) tensor ((13)C & (15)N) and heteronuclear correlation ((1)H-(13)C) data. The polymorph has two molecules per asymmetric unit (Z' = 2) and is thus the first conformational characterization with Z' > 1 established solely by SSNMR. Experimental data are correlated with structure through a series of computational models that extensively sample all conformations. For each computational model, corresponding tensor values are computed to supply comparisons with experimental information which, in turn, establishes paclitaxel's structure. Heteronuclear correlation data at thirteen key positions provide shift assignments to the asymmetric unit for each comparison. The two distinct molecules of the asymmetric unit possess nearly identical baccatin III moieties with matching conformations of the C10 acetyl moiety and, specifically, the torsion angle formed by C30-O-C10-C9. Additionally, both are found to exhibit an extended conformation of the phenylisoserine sidechain at C13 with notable differences in the dihedral angles centered around the rotation axes of O-C13, C2'-C1' and C3'-C2'.     

Automated compound verification using 2D-NMR HSQC data in an open-access environment

Since the introduction of NMR prediction software, medicinal chemists have imagined submitting their compounds to corporate compound registration systems that would ultimately display a simplified pass/fail result. We initially implemented such a system based on HPLC and liquid chromatography mass spectrometry (LCMS) data that is embedded within our industry standard sample submission and registration process. By using gradient-heteronuclear single quantum coherence (HSQC) experiments, we have extended this concept to NMR data through a comparison of experimentally acquired data against predicted (1)H and (13)C NMR data. Integration of our compound registration system with our analytical instruments now provides our chemists unattended and automated NMR verification for collections of submitted compounds. The benefits achieved from automated processing and interpretation of results produced enhanced confidence in our compound library and released the chemists from the tedium of manipulating large amounts of data. This allows scientists to focus more of their attention to the drug discovery process.     

Conformational preferences of the aglycon moiety in models and analogs of GlcNAc-Asn linkage: crystal structures and ab initio quantum chemical calculations of N-(beta-D-glycopyranosyl)haloacetamides

The biological addition of oligosaccharide structures to asparagine residues of N-glycoproteins influences the properties and bioactivities of these macromolecules. The linkage region constituents, 2-acetamino-2-deoxy-beta-D-glucopyranose monosaccharide (GlcNAc) and L-asparagine amino acid (Asn), are conserved in the N-glycoproteins of all eukaryotes. In order to gain information about the structure and dynamics of glycosylated proteins, two chloroacetamido sugars, Glc betaNAcNHCOCH2Cl and Man betaNHCOCH2Cl, have been synthesized, and their crystal structures have been solved. Structural comparison with a series of other models and analogs gives insight about the influence of the N-acetyl group at position C2 on the conformation of the glycan-peptide linkage at C1. Interestingly, this N-acetyl group also influences the packing and network of hydrogen bonds with involvement in weak hydrogen bonds C-H...X that are of biological importance. DFT ab initio calculations performed on a series of models and analogs also confirm that the GlcNAc derivatives present different preferred conformation about the N-CO-CH2-X (chi2) torsion angle of the glycan-peptide linkage, when compared to other monosaccharide derivatives. The energy profiles that have been obtained will be useful for parametrization of molecular mechanics force-field. The conjunction of crystallographic and computational chemistry studies provides arguments for the structural effect of the N-acetyl group at C2 in establishing an extended conformation that presents the oligosaccharide away from the protein surface.