Voronoi binding site models: Calculation of binding modes and influence of drug binding data accuracy

A new and accurate method for calculating the geometrically allowed modes of binding of a ligand molecule to a Voronoi site model is reported. It is shown that the feasibility of the binding of a group of atoms to a Voronoi site reduces to a simple set of linear and quadratic inequalities and quadratic equalities which can be solved by minimization of a simple function. Newton's numerical method of solution coupled to a line search proved to be successful. Moreover, we have developed efficient molecular and site data bases to discard quickly infeasible binding modes without time-consuming numerical calculation. The method is tested with a data set consisting of the binding constants for a series of biphenyls binding to prealbumin. After determination of the conformation space of the molecules and proposal of a Voronoi site geometry, the geometrically feasible modes are calculated and the energy interaction parameters determined to fit the observed binding energies to the site within experimental error ranges. We actually allowed these ranges to vary in order to study the influence of their broadness on the site geometry and found that as they increase, one can first model the receptor as a three-region site then as a single region site, but never as a two-region site.     

Voronoi binding site models

A frequently occurring problem in drug design and enzymology is that the binding constants for several compounds to the same site are known, but the geometry and energetic interactions of the site are not. This paper presents in detail a novel approach to the problem which accurately but compactly represents the allowed conformation space of each ligand, accurately depicts their three-dimensional structures, and realistically allows each ligand to adopt the conformation and positioning in the site which is most favorable energetically. The investigator supplies only the ligand structures and observed binding free energies, along with a proposed site geometry. With no further assumptions about how the ligands bind and what parts of the ligands are important in determining the binding, the algorithm fits the observed binding energies without leaving outliers, predicts exactly how each of the given ligands binds in the site, and predicts the strength and mode of binding of new compounds, regardless of chemical similarity to the original set of ligands. The method is illustrated by devising a simple site that accounts for the binding of five polychlorinated biphenyls to thyroxine binding prealbumin. This model then predicts the binding energies correctly for an additional six biphenyls, and fails on one compound.     

Intervals and the deduction of drug binding site models

In the search for new drugs, it often occurs that the binding affinities of several compounds to a common receptor macromolecule are known experimentally, but the structure of the receptor is not known. This article describes an extraordinarily objective computer algorithm for deducing the important geometric and energetic features of the common binding site, starting only from the chemical structures of the ligands and their observed binding. The user does not have to propose a pharmacophore, guess the bioactive conformations of the ligands, or suggest ways to superimpose the active compounds. The method takes into account conformational flexibility of the ligands, stereospecific binding, diverse or unrelated chemical structures, inaccurate or qualitative binding data, and the possibility that chemically similar ligands may or may not bind to the receptor in similar orientations. The resulting model can be viewed graphically and interpreted in terms of one or more binding regions of the receptor, each preferring to be occupied by various sorts of chemical groups. The model always fits the given data completely and can predict the binding of any other ligand, regardless of chemical structure. The method is an outgrowth of distance geometry and Voronoi polyhedra site modeling but incorporates several novel features. The geometry of the ligand molecules and the site is described in terms of intervals of internal distances. Determining the site model consists of reducing the uncertainty in the interregion distance intervals, and this uncertainty is described as intervals of intervals. Similarly, the given binding affinities and their experimental uncertainties are treated as intervals in the affinity scale. The final site model specifies an entire region of interaction energy parameters that satisfy the training set rather than a single set of parameters. Predicted binding for test compounds results in an interval which, when compared to the experimental interval, may be correct, incorrect, or vague. There is a pervasive ternary logic involved in the assessment of predictions, in the search for a satisfactory model, and in judging whether a given molecule may bind in a particular orientation: true, false, or maybe. The approach is illustrated on an extremely simple artificial example and on a real data set of cocaine analogues binding to a nerve membrane receptor in vitro. © 1995 by John Wiley & Sons, Inc.     

A cation-pi binding interaction with a tyrosine in the binding site of the GABAC receptor

GABA(C) (rho) receptors are members of the Cys-loop superfamily of neurotransmitter receptors, which includes nicotinic acetylcholine (nACh), 5-HT(3), and glycine receptors. As in other members of this family, the agonist binding site of GABA(C) receptors is rich in aromatic amino acids, but while other receptors bind agonist through a cation-pi interaction to a tryptophan, the GABA(C) binding site has tyrosine at the aligning positions. Incorporating a series of tyrosine derivatives at position 198 using unnatural amino acid mutagenesis reveals a clear correlation between the cation-pi binding ability of the side chain and EC(50) for receptor activation, thus demonstrating a cation-pi interaction between a tyrosine side chain and a neurotransmitter. Comparisons among four homologous receptors show variations in cation-pi binding energies that reflect the nature of the cationic center of the agonist.     

Geometric accuracy of three-dimensional molecular overlays

This study examines the dependence of molecular alignment accuracy on a variety of factors including the choice of molecular template, alignment method, conformational flexibility, and type of protein target. We used eight test systems for which X-ray data on 145 ligand-protein complexes were available. The use of X-ray structures allowed an unambiguous assignment of bioactive overlays for each compound set. The alignment accuracy depended on multiple factors and ranged from 6% for flexible overlays to 73% for X-ray rigid overlays, when the conformation of the template ligand came from X-ray structures. The dependence of the overlay accuracy on the choice of templates and molecules to be aligned was found to be the most significant factor in six and seven of the eight ligand-protein complex data sets, respectively. While finding little preference for the overlay method, we observed that the introduction of molecule flexibility resulted in a decrease of overlay accuracy in 50% of the cases. We derived rules to maximize the accuracy of alignment, leading to a more than 2-fold improvement in accuracy (from 19% to 48%). The rules also allowed the identification of compounds with a low (<5%) chance to be correctly aligned. Last, the accuracy of the alignment derived without any utilization of X-ray conformers varied from <1% for the human immunodeficiency virus data set to 53% for the trypsin data set. We found that the accuracy was directly proportional to the product of the overlay accuracy from the templates in their bioactive conformations and the chance of obtaining the correct bioactive conformation of the templates. This study generates a much needed benchmark for the expectations of molecular alignment accuracy and shows appropriate usages and best practices to maximize hypothesis generation success.     

Enhancing the accuracy of virtual screening: molecular dynamics with quantum-refined force fields

Summary A methodology aimed at improving the accuracy of current docking–scoring procedures is proposed, and validated through detailed tests of its performance in predicting the activity of HIV-1 protease inhibitors. This methodology is based on molecular dynamics simulations using a force field whose effective charges are refined by means of a novel procedure that relies on quantum-mechanical calculations and preserves the internal consistency of the parameterization scheme.     

CoMFA-based comparison of two models of binding site on adenosine A1 receptor

A set of 32 N6-substituted adenosines and 22 8-substituted xanthines with affinity for adenosine A1 receptors was subjected to three-dimensional quantitative structure-affinity relationship analysis using comparative molecular field analysis (CoMFA). The aim was to compare two modes of binding to the receptor – `N6-C8' and `N6-N7'. Good models with high predictive power and stability were obtained. A comparison of these models gives the following results: (a) Inclusion of both steric and electrostatic fields in CoMFA generates better predictive models compared to models based on steric or electrostatic fields alone. (b) The `N6-N7' CoMFA models are slightly better than the `N6-C8' ones. (c) Steric restriction exists around the N6-H in the `N6-N7' steric field map, which is absent in the `N6-C8' steric field map. This report demonstrates that the `N6-N7' mode of binding is a further development of the `N6-C8' model with a slightly better predictive ability and more accurate steric and electrostatic overlaps between agonists and antagonists.     

Enhancing the accuracy of measurement of small molecule organic biomarkers

Analytical and Bioanalytical Chemistry - Over two decades, the Organic Analysis Working Group (OAWG) of the Consultative Committee for Amount of Substance: Metrology in Chemistry and Biology (CCQM)...     

Enhancing the selectivity of an iron binding hydrogel

Iron overload is a critical clinical condition that can be controlled by diet and the use of iron-specific chelating agents. An effective oral formulation of an iron chelator should be nontoxic and selective toward iron while maintaining high affinity for iron. In this study, hydrogels containing 2,3 dihydroxybenzoic acid (2,3 DHBA), a portion of the metal chelating domain of enterobactin, were synthesized as a potential non-absorbed chelator for iron in the gastrointestinal tract. A series of polymeric chelators with various hydrogel:DHBA ratios were prepared. The iron-binding properties of the hydrogels were found to depend on the concentration of 2,3 DHBA groups on polymer chains. A 0.005 ratio of PAAm:DHBA was found to have an optimum affinity (log K = 27.01), selectivity, and high binding capacity for Fe(III).     

Synthetic models for the transition metal binding site of bleomycin. Remarkable improvement of dioxygen activating capability

Model compounds for the metal binding site of bleomycin with a CH₂ CONH₂ group (PYML-3) or atert-butyl group (PYML-4) as steric environmental factors are synthesized. These exhibit metal binding properties similar to those of bleomycin. In particular, PYML-4 shows oxygen activation up to 71 % of that of bleomycin.     

Ligand design by targeting a binding site water

Solvent reorganization is a major driving force of protein–ligand association, but the contribution of binding site waters to ligand affinity is poorly understood. We investigated how altered interactions with a water network can influence ligand binding to a receptor. A series of ligands of the A_2A adenosine receptor, which either interacted with or displaced an ordered binding site water, were studied experimentally and by molecular dynamics simulations. An analog of the endogenous ligand that was unable to hydrogen bond to the ordered water lost affinity and this activity cliff was captured by molecular dynamics simulations. Two compounds designed to displace the ordered water from the binding site were then synthesized and evaluated experimentally, leading to the discovery of an A_2A agonist with nanomolar activity. Calculation of the thermodynamic profiles resulting from introducing substituents that interacted with or displaced the ordered water showed that the gain of binding affinity was enthalpy driven. Detailed analysis of the energetics and binding site hydration networks revealed that the enthalpy change was governed by contributions that are commonly neglected in structure-based drug optimization. In particular, simulations suggested that displacement of water from a binding site to the bulk solvent can lead to large energy contributions. Our findings provide insights into the molecular driving forces of protein–ligand binding and strategies for rational drug design.

Solvent reorganization is a major driving force of protein–ligand association, but the contribution of binding site waters to ligand affinity is poorly understood.     

A ligand-based approach to mining the chemogenomic space of drugs

The practical implementation and validation of a ligand-based approach to mining the chemogenomic space of drugs is presented and applied to the in silico target profiling of 767 drugs against 684 targets of therapeutic relevance. The results reveal that drugs targeting aminergic G protein-coupled receptors (GPCRs) show the most promiscuous pharmacological profiles. The detection of cross-pharmacologies between aminergic GPCRs and the opioid, sigma, NMDA, and 5-HT3 receptors aggravate the potential promiscuity of those drugs, predominantly including analgesics, antidepressants, and antipsychotics.     

Determination of binding site concentration and average binding constant by the method of equilibrium sorption: Evidence for lysine as a principal receptor site for collagen-enzyme binding

The contribution of lysyl ϵ-amino groups to the noncovalent binding on enzymes to collagen has been determined by the method of equilibrium sorption. The sorption or binding of enzyme by chemically modified, and by the corresponding unmodified membraneous collagen, was determined as a function of enzyme concentration. Chemical modification of lysyl ϵ-amino groups was accomplished by carbamylation of the collagen membrane with potassium cyanate. The collagen membranes were contacted with a purified β-galactosidase (E. coli K₁₂). The enzyme was purified by affinity chromato- graphy. Analysis of the resultant sorption isotherms allowed determination of the heterogeneity index (a) and the concentration of active binding sites (A_o). Binding constants (K_o) and the free energy of binding (δG) were also computed from the sorption data.The results of these studies have shown that lysyl ϵ-amino groups of collagen are primary receptor sites for enzyme binding (β-galactosidase, E. coli K₁₂) and that carbamylation of the collagen membrane resulted in a decrease in active binding sites with little or no change in the binding constant of the remaining sites.     

Investigation of the copper binding site and the role of histidine as a ligand in riboflavin binding protein

Riboflavin Binding Protein (RBP) binds copper in a 1:1 molar ratio, forming a distinct well-ordered type II site. The nature of this site has been examined using X-ray absorption and pulsed electron paramagnetic resonance (EPR) spectroscopies, revealing a four coordinate oxygen/nitrogen rich environment. On the basis of analysis of the Cambridge Structural Database, the average protein bound copper-ligand bond length of 1.96 A, obtained by extended x-ray absorption fine structure (EXAFS), is consistent with four coordinate Cu(I) and Cu(II) models that utilize mixed oxygen and nitrogen ligand distributions. These data suggest a Cu-O 3N coordination state for copper bound to RBP. While pulsed EPR studies including hyperfine sublevel correlation spectroscopy and electron nuclear double resonance show clear spectroscopic evidence for a histidine bound to the copper, inclusion of a histidine in the EXAFS simulation did not lead to any significant improvement in the fit.     

Multiple flexible alignment with SEAL: a study of molecules acting on the colchicine binding site

An extension of the steric and electrostatic alignment alignment (SEAL) method (MultiSEAL) is described that allows the overlay of multiple molecules and conformations. The method is well-suited for the systematic study of possible alignments, also revealing information about the conformational energies associated with a given overlay. It has been tested on three examples: angiotensin II antagonists, 5-HT3 antagonists, and dopaminergic compounds. The utility of the method is further demonstrated in an analysis of molecules that putatively bind to the colchicine site of tubulin. On the basis of its overlay with colchicine, allocolchicine, 2-methoxy-5-(2',3',4'-trimethoxyphenyl)tropone, and combretastatin A-4, it appears that 2-methoxyestradiol (2-ME) is unlikely to fit the colchine site properly. The weak antimitotic activity of 2-ME may be explained by its partial fit in the site.     

Thermodynamic analysis of a hydrophobic binding site: probing the PDZ domain with nonproteinogenic peptide ligands

[structure: see text] Isothermal titration calorimetry (ITC) is used to study the thermodynamic consequences of systematically modifying the hydrophobic character of a single residue in a series of protein-binding ligands. By substituting standard and nonproteinogenic aliphatic amino acids for the C-terminal valine of the hexapeptide KKETEV, binding to the third PDZ domain (PDZ3) of the PSD-95 protein is characterized by distinct changes in the Gibbs free energy (DeltaG), enthalpy (DeltaH), and entropy (TDeltaS) parameters. One notable observation is that peptide binding affinity can be improved with a nonstandard residue.