The measurement of molecular diversity: A three-dimensional approach

Summary This paper describes a method for selecting a small, highly diverse subset from a large pool of molecules. The method has been employed in the design of combinatorial synthetic libraries for use in high-throughput screening for pharmaceutical lead generation. It computes diversity in terms of the main factors relevant to ligand-protein binding, namely the three-dimensional arrangement of steric bulk and of polar functionalities and molecular entropy. The method was used to select a set of 20 carboxylates suitable for use as side-chain precursors in a polyamine-based library. The method depends on estimates of various physical-chemical parameters involved in ligand-protein binding; experiments examined the sensitivity of the method to these parameters. This paper compares the diversity of randomly and rationally selected side-chain sets; the results suggest that careful design of synthetic combinatorial libraries may increase their effectiveness several-fold.     

Deciphering common failures in molecular docking of ligand-protein complexes

Common failures in predicting crystal structures of ligand-protein complexes are investigated for three ligand-protein systems by a combined thermodynamic and kinetic analysis of the binding energy landscapes. Misdocked predictions in ligand-protein docking are classified as `soft' and `hard' failures. While a soft failure arises when the search algorithm is unable to find the global energy minimum corresponding to the crystal structure, a hard failure results from a flaw of the energy function to qualify the crystal structure as the predicted lowest energy conformation in docking simulations. We find that neither the determination of a single structure with the lowest energy nor finding the most common binding mode is sufficient to predict crystal structures of the complexes, which belong to the category of hard failures. In a proposed hierarchical approach, structural similarity clustering of the conformations, generated from equilibrium simulations with the simplified energy function, is followed by energy refinement with the AMBER force field. This protocol, that involves a hierarchy of energy functions, resolves some common failures in ligand-protein docking and detects crystallographic binding modes that were not found during docking simulations.     

Clean STD-NMR spectrum for improved detection of ligand-protein interactions at low concentration of protein

Saturation transfer difference (STD)-NMR has been widely used to screen ligand compound libraries for their binding activities to proteins and to determine the binding epitopes of the ligands. We report herein, a Clean STD-NMR method developed to overcome false positives (artifacts) observed in the STD-NMR spectrum due to the power spillover of RF irradiation. The method achieved higher degree of resonance saturation through digital editing of two STD-NMR spectra to generate a concatenated difference spectrum and three times of sensitivity enhancement for a loose binding complex involving DNA oligonucleotide and an RNA-binding protein, CUGBP-1ab (25.2 kDa). The interesting binding characteristics of the complex dCTGTCT-CUGBP1ab were obtained. The method was applied to a mixture of small ligand and bovine serum albumin protein (BSA, 66.3 kDa), and detected the intermolecular contacts at a BSA concentration as low as 0.1 μM, a working concentration useful for the detection of proteins of low solubility at biologically relevant conditions.     

Evaluating docked complexes with the HINT exponential function and empirical atomic hydrophobicities

Summary Methods that predict geometries of ligands binding to receptor molecules can facilitate ligand discovery and yield information on the factors governing complementarity. Here, the use of atomic hydrophobicities in evaluating binding modes has been examined with four ligand-receptor complexes of known structure. In each system, hundreds of hypothetical binding orientations were generated with DOCK and evaluated using the HINT (Hydropathic INTeractions) exponential function and atomic hydrophobic constants. In three of the four systems, the experimental binding mode received the best HINT score; in the fourth system, the experimental binding mode scored only slightly lower than a similar, apparently reasonable orientation. The HINT function may be generally useful as a scoring method in molecular docking.     

Molecular dynamics simulation study on the interaction of KRN 7000 and three analogues with human CD1d

Many analogues of KRN 7000, a synthetic glycolipid (α-galactosylceramide) exhibiting immuno-stimulatory activity and antitumor properties, were previously synthesized and tested in order to understand the reasons for the resulting biological activity and Th1/Th2 cytokine profile. Principles have been established for the interaction of such glycolipids with the human CD1d molecule but the exact mechanism by which different ligands with the same polar head elicit distinct biological responses remains unclear. Based on these experiments and on the available crystal structures, protein-ligand interactions are explored using molecular dynamics simulations. Hydrogen bond interactions are examined with regard to the polar group orientation. The influence of modulations on the dynamic behavior of the CD1d-glycolipid complex is addressed. Overall, our data support the mechanism by which the shortening of the α-GalCer sphingosine chain causes a significant twist of the CD1d α1 helix structure from residue Phe84 that affects the position of CD1d residues involved in the TCR recognition.     

A molecular model for the active site of S-adenosyl-l-homocysteine hydrolase

Summary S-adenosyl-l-homocysteine hydrolase (AdoHcy hydrolase, EC 3.3.1.1.), a specific target for antiviral drug design, catalyzes the hydrolysis of AdoHcy to adenosine (Ado) and homocysteine (Hcy) as well as the synthesis of AdoHcy from Ado and Hcy. The enzyme isolated from different sources has been shown to contain tightly bound NAD⁺.Based on the 2.0 Å-resolution X-ray crystal structure of dogfish lactate dehydrogenase (LDH), which is functionally homologous to AdoHcy hydrolase, and the primary sequence of rat liver AdoHcy hydrolase, we have derived a molecular model of an extended active site for AdoHcy hydrolase. The computational mutation was performed using the software MUTAR (Yeh et al., University of Kansas, Lawrence), followed by molecular mechanics optimizations using the programs AMBER (Singh et al., University of California, San Francisco) and YETI (Vedani, University of Kansas). Solvation of the model structure was achieved by use of the program SOLVGEN (Jacober, University of Kansas); 56 water molecules were explicitly included in all refinements. Some of these may be involved in the catalytic reaction.We also studied a model of the complex of AdoHcy hydrolase with NAD⁺, as well as the ternary complexes of the redox reaction catalyzed by AdoHcy hydrolase and has been used to differentiate the relative binding strength of inhibitors.     

Molecular recognition studies with a simple dipyrrinone

We present our investigations of 2-ethyl-3-methyl-(10H)-dipyrrin-1-one, its self-association, and anion binding properties. This receptor is easily accessible in a facile single step synthesis with a straightforward workup. An examination of the concentration dependence of the dipyrrinone NH chemical shifts in CDCl₃ and (CDCl₂)₂ over the temperature range from −20 °C to 100 °C determined the self-association constant to be 3850 M⁻¹. Molecular recognition studies have shown that it has a preference for guests with an OH moiety, such as hydrogen sulfate (HSO₄⁻) and carboxylic acids (RCO₂H).     

Probing ligand-protein recognition with sum-frequency generation spectroscopy: the avidin-biocytin case.

Infrared/visible sum-frequency generation (SFG) spectroscopy is used to study the recognition of a protein (avidin) by a derived vitamin (biocytin) adsorbed on a calcium fluoride substrate. The specificity of the process is tested by replacing avidin with bovine serum albumin or presaturated avidin. The SFG spectroscopy shows drastic modifications in the CH and NH spectral ranges only upon exposure of the biocytin film to avidin. The comparison of the SFG data with Fourier transform infrared reflection absorption spectra (FT-IRRAS) in the same spectral ranges illustrates the advantages of nonlinear spectroscopy for studying and detecting recognition between biomolecules.     

Caffeine interactions with nucleic acids. Molecular mechanics calculations of model systems for explanation of mechanisms of biological actions

To understand the molecular mechanisms of the influence of caffeine (CAF) on DNA functioning, molecular mechanics calculations of the interaction energy of CAF with nucleic acid bases and base pairs have been performed. The calculations reveal three types of mutual CAF–base (and CAF–base pair) arrangements corresponding to minima of the interaction energy. Besides well-known stacking mutual positions of the molecules, two other types of arrangements are revealed and studied. One of these arrangements corresponds to the nearly in-plane position of CAF and base (or base pair) and the formation of a single hydrogen bond. Another type of minimum corresponds to nearly perpendicular arrangements of the molecular planes and the formation of intermolecular hydrogen bonds. These two arrangements are possible both for individual nucleic acid monomers and for DNA duplexes. The calculations suggest the molecular mechanisms of the influence of CAF on DNA interactions with other biologically active molecules.From the Proceedings of the 28th Congreso de Químicos Teóricos de Expresión Latina (QUITEL 20002).     

Evidence for strong heterodimeric interactions of phenylboronic acids with amino acids

Strong heterodimeric interactions of phenylboronic acids with l-proline or betaine are evident in the solid state. The interaction energy is over 23 kcal/mol (at MP2/6-31+G^∗).     

Compass: A shape-based machine learning tool for drug design

Summary Building predictive models for iterative drug design in the absence of a known target protein structure is an important challenge. We present a novel technique, Compass, that removes a major obstacle to accurate prediction by automatically selecting conformations and alignments of molecules without the benefit of a characterized active site. The technique combines explicit representation of molecular shape with neural network learning methods to produce highly predictive models, even across chemically distinct classes of molecules. We apply the method to predicting human perception of musk odor and show how the resulting models can provide graphical guidance for chemical modifications.     

Molecularly imprinted supermacroporous cryogels for cytochrome c recognition

Molecular imprinting is an attractive biomimetic approach that creates specific recognition sites for the shape and functional group arrangement to template molecules. The purpose of this study is to prepare cytochrome c-imprinted poly(hydroxyethyl methacrylate) (PHEMA)-based supermacroporous cryogel which can be used for the separation of cytochrome c from protein mixtures. N-Methacryloyl-(L)-histidinemethylester (MAH) was used as the metal-coordinating monomer. In the first step, Cu(2+) was complexed with MAH, and the cytochrome c imprinted PHEMA (MIP) cryogel was prepared by free radical cryopolymerization initiated by N,N,N',N'-tetramethylene diamine at -12°C. After polymerization is completed, the template cytochrome c molecules were removed from the MIP cryogel using 0.5 M NaCl solution. The maximum cytochrome c binding amount was 126 mg/g polymer. Selective binding studies were performed in the presence of lysozyme and bovine serum albumin. The relative selectivity coefficients of MIP cryogel for cytochrome c/lysozyme and cytochrome c/bovine serum albumin were 1.7 and 5.2 times greater than those of the non-imprinted PHEMA cryogel, respectively. The selectivity of MIP cryogel for cytochrome c was also confirmed with fast protein liquid chromatography. The MIP cryogel could be used many times with no remarkable decrease in cytochrome c binding capacity.     

Fluorescence sensing of tartaric acid: a case of excimer emission caused by hydrogen bond-mediated complexation

A novel quinoline based receptor that shows monomer emission quenching followed by intramolecular excimer emission upon hydrogen bond mediated complexation of tartaric acid has been designed and synthesized. The excimer emission has been used to confirm the selective recognition of tartaric acid over its nonhydroxy analogue, succinic acid. Binding ability was studied by ¹H NMR, UV-vis and fluorescence spectroscopic methods.     

DNA小沟结合二脒:水分子的识别作用

采用分子动力学模拟了DB921-DNA复合物, 通过7 ns的模拟研究表明: DB921一端的氨基氮原子与一个水分子形成氢键, 同时, 水分子又与DNA的5位A碱基的氮原子形成一个氢键. 水分子在DB921与DNA小沟结合中起了桥连的作用, 使得直线型的芳香二脒化合物DB921通过水桥与DNA小沟结合, 水分子诱导DB921分子与DNA的小沟域构型相适应, 与DNA小沟域的AATTC碱基有较强的结合作用. 在分子水平上提供了DB921与双螺旋DNA相互作用的结构及复合物的动态变化情况, 指出水分子在DNA小沟结合二脒化合物中的识别作用, 为设计出更高生物活性的DNA小沟结合剂提供一定的理论依据.     

Tautomerism of xanthine and alloxanthine: A model for substrate recognition by xanthine oxidase

Summary Tautomerism of neutral xanthine and alloxanthine has been examined both in the gas phase and in aqueous solution. The tautomeric preference in the gas phase has been studied by means of semiempirical and ab initio quantum-mechanical computations with inclusion of correlation effects at the Møller-Plesset level, and from density-functional calculations. The influence of solvent on the relative stability between tautomers has been estimated from self-consistent reaction field calculations performed with different models. The results provide a detailed picture of tautomerism for these biologically relevant purine bases. The functional implications in the recognition by xanthine oxidase are analyzed from inspection of the interaction patterns of the most stable tautomeric forms. A model for the recognition of these purine derivatives in the enzyme binding site is discussed.     

一种新型芳香二胺桥联β-环糊精对染料的分子识别

以单-(6-对甲苯磺酰基)-β-环糊精和3,3′-亚甲基联苯胺为原料, 合成了一种新型刚性结构的芳香二胺桥联环糊精, 3,3′-亚甲基联苯胺桥联(6-氨基-6-脱氧-β-环糊精)2. 并用荧光光谱和紫外光谱技术分别测定了在25 ℃时, pH为7.20的磷酸缓冲溶液中β-环糊精(1)和新型桥联环糊精(2)与几种染料分子, 如吖定红(AR)、中性红(NR)、2-对甲苯胺基-6-萘磺酸钠(TNS)、1-苯胺基-8-萘磺酸铵(ANS)、罗丹明B(RhB)和亮绿(BG)形成超分子配合物的稳定常数KS. 化学计量比表明, 桥联环糊精2与客体形成了1∶1的超分子配合物, 其对客体的键合能力和分子选择性远远强于母体?茁-环糊精, 如桥联环糊精2对BG的键合能力可以达到母体环糊精的22.2倍. 从主-客体间的尺寸匹配关系和多重识别机理等方面探讨了桥联环糊精对客体分子的协同键合作用.     

Asymmetric enolate alkylation via templation with chiral synthetic receptors

C₃-Symmetric chiral receptors have been developed for enantioselective alkylation of sodium enolates of active methylene compounds. It has been demonstrated that a 1:1 binding complex forms between these receptors and sodium enolates in THF-d₈/CD₃CN by ¹H NMR titration experiments. Moderate enantiomeric enrichment of the benzylation product of 2-acetylcyclohexanone has been demonstrated using this strategy. Templation of enolate alkylation by synthetic receptors represents a new approach to asymmetric induction.     

Fluorophore-linked zinc(II)dipicolylamine coordination complexes as sensors for phosphatidylserine-containing membranes

A series of Zn²⁺-2,2′-dipicolylamine (Zn²⁺-DPA) coordination complexes with an attached NBD fluorophore are synthesized and evaluated as fluorescent sensors. The sensors do not respond to vesicles composed of zwitterionic phosphatidylcholine, but the NBD fluorescence emission is enhanced in the presence of anionic vesicles. A sensor with two Zn²⁺-DPA units and a hydrophilic tris(ethyleneoxy) linker produced a larger emission enhancement than an analogue with a butyl linker, and titration with 1:1 POPC:POPS vesicles lead to an apparent phospholipid association constant of 5.3×10⁴ M⁻¹. The sensor can detect the presence of vesicles containing as little as 5% phosphatidylserine. The sensing effect apparently requires a membrane surface because the sensors do not respond to a phosphatidylserine derivative that is monodispersed in aqueous solution.