Gold(I) and mercury(II)-isoelectronic ions with strongly different chemistry: ab initio QMCF molecular dynamics simulations of their hydration structure.

The hydration structure of the isoelectronic Au(I) and Hg(II) ions was determined by means of ab initio quantum mechanical charge field molecular dynamics (QMCF MD) simulations. The two hydrates proved as very labile but entirely different in their structural features. While Hg(II) forms two distinct hydration shells, Au(I) is characterized by an additional extended first shell (meso-shell) which has a considerable influence on all data extracted from the simulation trajectory, namely, radial and angular distribution functions, coordination number distribution, and dynamical data such as mean ligand residence times (MRT) and vibrational frequencies. The short MRT values of the first shell ligands, amounting to a few picoseconds, lead to the simultaneous presence of a number of hydrate complexes with differing geometries, which explains the difficulties in assigning structural data to spectroscopic measurements. The results presented here demonstrate that isoelectronic transition metal ions can show strongly different chemical properties, which cannot be explained on the basis of their different charge alone. The importance of including the second hydration shell and thus the intershell hydrogen bonds in the quantum mechanical treatment of the simulation is clearly proven.

Preferential amino acid sequences in alumina-catalyzed peptide bond formation.

The catalytic effect of activated alumina on amino acid condensation was investigated. The readiness of amino acids to form peptide sequences was estimated on the basis of the yield of dipeptides and was found to decrease in the order glycine (Gly), alanine (Ala), leucine (Leu), valine (Val), proline (Pro). For example, approximately 15% Gly was converted to the dipeptide (Gly(2)), 5% to cyclic anhydride (cyc(Gly(2))) and small amounts of tri- (Gly(3)) and tetrapeptide (Gly(4)) were formed after 28 days. On the other hand, only trace amounts of Pro(2) were formed from proline under the same conditions. Preferential formation of certain sequences was observed in the mixed reaction systems containing two amino acids. For example, almost ten times more Gly-Val than Val-Gly was formed in the Gly+Val reaction system. The preferred sequences can be explained on the basis of an inductive effect that side groups have on the nucleophilicity and electrophilicity, respectively, of the amino and carboxyl groups. A comparison with published data of amino acid reactions in other reaction systems revealed that the main trends of preferential sequence formation were the same as those described for the salt-induced peptide formation (SIPF) reaction. The results of this work and other previously published papers show that alumina and related mineral surfaces might have played a crucial role in the prebiotic formation of the first peptides on the primitive earth.

Activated alumina as an energy source for peptide bond formation: consequences for mineral-mediated prebiotic processes.

The catalytic properties of various forms of alumina were tested for alanine dimerization reaction. The catalytic efficiency of alumina depends on the structure, as well as on acid/base properties of the catalyst. The highest yields of Ala2 were achieved on activated alumina with surface of neutral pH (about 3% conversion after 2 weeks). Thermal analysis of Ala + alumina reaction systems shows that the thermal behavior of amino acid changes substantially in contact with the activated surface of the alumina catalyst. The reaction of Ala is detected as being strongly endothermic by differential thermal analysis of pure amino acid (above 250 degrees C). The alanine endothermic reaction is shifted substantially to lower values (below 200 degrees C) and hardly detectable if activated alumina is present. The reaction mechanism of amino acid activation on alumina surface and its significance for mineral-catalyzed prebiotic peptide bond formation are discussed.

Elbow flexibility and ligand-induced domain rearrangements in antibody Fab NC6.8: large effects of a small hapten.

Four 700-ps molecular dynamics simulations were carried out to analyze the structural dynamics of the antigen-binding antibody fragment NC6.8, which is known to exhibit large structural changes upon complexation. The first simulation was started from the x-ray structure of the uncomplexed Fab and produced trajectory averages that closely match the crystallographic results. It allowed assessment of the flexibility of the Fab, revealing an elbow motion of the variable domains with respect to the constant domains. The second simulation was started from the uncomplexed x-ray structure after insertion of the ligand into the binding site. This perturbation resulted in a significantly altered trajectory, with quaternary structural changes corresponding in many aspects to the experimental differences between complexed and uncomplexed state. The observed trend toward a smaller elbow angle and a higher flexion of the H-chain could also be seen in the third simulation, which was started from the x-ray structure of the complex. The changes were revealed to be a clear consequence of the complexation with the ligand because in the fourth simulation (started from the experimental complex structure after removal of the hapten) the Fab remained close to its initial structure. Analyses of the quaternary structure and the binding site of Fab NC6.8 are presented for all four simulations, and possible interpretations are discussed.

Automated docking of ligands to antibodies: methods and applications.

Many approaches to studying protein-ligand interactions by computational docking are currently available. Given the structures of a protein and a ligand, the ultimate goal of all docking methods is to predict the structure of the resulting complex. This requires a suitable representation of molecular structures and properties, search algorithms to efficiently scan the configuration space for favorable interaction geometries, and accurate scoring functions to evaluate and rank the generated orientations. For many of the available methods, tests on experimentally known antibody-antigen or antibody-hapten complexes have appeared in the literature. In addition, some of them have been used in predictive studies on antibody-ligand interactions to provide structural insights where adequate experimental information is missing. The AutoDock program is presented as example of a method for flexibly docking ligands to antibodies. Applying parameters of the second-generation AMBER force field, three antibody-hapten complexes (AN02, DB3, NC6.8) are used as new test cases to analyze the ability of the method to reproduce experimental findings. The X-ray structures could be reconstituted and the corresponding solutions were ranked with best energy score in all cases. Docking to the free instead of the complexed NC6.8 structure indicated the limits of the rigid protein treatment, although fairly good guesses about the location of the binding site and the contact residues could still be obtained if conformational flexibility was allowed at least in the ligand.

Application of multivariate data analysis methods to comparative molecular field analysis (CoMFA) data: proton affinities and pKa prediction for nucleic acids components.

Multivariate data analysis methods (Principal Component Analysis (PCA) and Partial Least Squares (PLS)) are applied to the analysis of the CoMFA (Comparative Molecular Field Analysis) data for several nucleic acids components. The data set includes nitrogenated bases, nucleosides, linear nucleotides, 3', 5'-cyclic nucleotides and oligonucleotides. PCA is applied to study the structure of the CoMFA data and to detect possible outliers in the data set. PLS is applied to correlate the CoMFA data with either calculated AM1 proton affinities or with experimental pKa values. The possibility of making a prediction of pKa values directly from 3D structures of the monomers for polynucleotides is also shown. The influence of the superposition criteria and of conformational changes along the glycosidic bond on the pKa prediction are studied as well.

Silica, alumina and clay catalyzed peptide bond formation: enhanced efficiency of alumina catalyst.

Catalytic efficiencies of clay (hectorite), silica and alumina were tested in peptide bond formation reactions of glycine (Gly), alanine (Ala), proline (Pro), valine (Val) and leucine (Leu). The reactions were performed as drying/wetting (hectorite) and temperature fluctuation (silica and alumina) experiments at 85 degrees C. The reactivity of amino acids decreased in order Gly > Ala > Pro approximately Val approximately Leu. The highest catalytic efficiency was observed for alumina, the only catalyst producing oligopeptides in all investigated reaction systems. The peptide bond formation on alumina is probably catalyzed by the same sites and via similar reaction mechanisms as some alumina-catalyzed dehydration reactions used in industrial chemistry.

Mutual amino acid catalysis in salt-induced peptide formation supports this mechanism's role in prebiotic peptide evolution.

The presence of some amino acids and dipeptides under the conditions of the salt-induced peptide formation reaction (aqueous solution at 85 degrees C, Cu(II) and NaCl) has been found to catalyze the formation of homopeptides of other amino acids, which are otherwise produced only in traces or not at all by this reaction. The condensation of Val, Leu and Lys to form their homodipeptides can occur to a considerable extent due to catalytic effects of other amino acids and related compounds, among which glycine, histidine, diglycine and diketopiperazine exhibit the most remarkable activity. These findings also lead to a modification of the table of amino acid sequences preferentially formed by the salt-induced peptide formation (SIPF) reaction, previously used for a comparison with the sequence preferences in membrane proteins of primitive organisms.

Peptides and the origin of life.

Considering the state-of-the-art views of the geochemical conditions of the primitive earth, it seems most likely that peptides were produced ahead of all other oligomer precursors of biomolecules. Among all the reactions proposed so far for the formation of peptides under primordial earth conditions, the salt-induced peptide formation reaction in connection with adsorption processes on clay minerals would appear to be the simplest and most universal mechanism known to date. The properties of this reaction greatly favor the formation of biologically relevant peptides within a wide variation of environmental conditions such as temperature, pH, and the presence of inorganic compounds. The reaction-inherent preferences of certain peptide linkages make the argument of 'statistical impossibility' of the evolutionary formation of the 'right' peptides and proteins rather insignificant. Indeed, the fact that these sequences are reflected in the preferential sequences of membrane proteins of archaebacteria and prokaryonta distinctly indicates the relevance of this reaction for chemical peptide evolution. On the basis of these results and the recent findings of self-replicating peptides, some ideas have been developed as to the first steps leading to life on earth.

The combination of salt induced peptide formation reaction and clay catalysis: a way to higher peptides under primitive earth conditions.

Two reactions with suggested prebiotic relevance for peptide evolution, the salt induced peptide formation reaction and the peptide chain elongation/stabilization on clay minerals have been combined in experimental series starting from dipeptides and dipeptide/amino acid mixtures. The results show that both reactions can take place simultaneously in the same reaction environment and that the presence of mineral catalysts favours the formation of higher oligopeptides. These findings lend further support to the relevance of these reactions for peptide evolution on the primitive earth. The detailed effects of the specific clay mineral depend both on the nature of the mineral and the reactants in solution.

Ligand binding by antibody IgE Lb4: assessment of binding site preferences using microcalorimetry, docking, and free energy simulations.

Antibody IgE Lb4 interacts favorably with a large number of different compounds. To improve the current understanding of the structural basis of this vast cross-reactivity, the binding of three dinitrophenyl (DNP) amino acids (DNP-alanine, DNP-glycine, and DNP-serine) is investigated in detail by means of docking and molecular dynamics free energy simulations. Experimental binding energies obtained by isothermal titration microcalorimetry are used to judge the results of the computational studies. For all three ligands, the docking procedure proposes two plausible subsites within the binding region formed by the antibody CDR loops. By subsequent molecular dynamics simulations and calculations of relative free energies of binding, one of these subsites, a tyrosine-surrounded pocket, is revealed as the preferred point of complexation. For this subsite, results consistent with experimental observations are obtained; DNP-glycine is found to bind better than DNP-serine, and this, in turn, is found to bind better than DNP-alanine. The suggested binding mode makes it possible to explain both the moderate binding affinity and the differences in binding energy among the three ligands.

Are prions a relic of an early stage of peptide evolution?

The rather unique properties of prions and their presence in very different kinds of living species suggest that this type of molecule was created at a very early stage of evolution and may even represent a relic from a time where peptide evolution was ongoing and RNA/DNA did not yet exist. A comparison of the most frequently occurring amino acid sequences in known prions with the sequences preferentially formed in the salt-induced peptide formation reaction, the most simple mechanism enabling the formation of peptides under primitive earth conditions, shows a remarkable coincidence that strongly supports this hypothesis.

Comparative molecular field analysis of artemisinin derivatives: ab initio versus semiempirical optimized structures.

Based on the belief that structural optimization methods, producing structures more closely to the experimental ones, should give better, i.e. more relevant, steric fields and hence more predictive CoMFA models, comparative molecular field analyses of artemisinin derivatives were performed based on semiempirical AM1 and HF/3-21G optimized geometries. Using these optimized geometries, the CoMFA results derived from the HF/3-21G method are found to be usually but not drastically better than those from AM1. Additional calculations were performed to investigate the electrostatic field difference using the Gasteiger and Marsili charges, the electrostatic potential fit charges at the AM1 level, and the natural population analysis charges at the HF/3-21G level of theory. For the HF/3-21G optimized structures no difference in predictability was observed, whereas for AM1 optimized structures such differences were found. Interestingly, if ionic compounds are omitted, differences between the various HF/3-21G optimized structure models using these electrostatic fields were found.

Ligand-induced domain movement in an antibody Fab: molecular dynamics studies confirm the unique domain movement observed experimentally for Fab NC6.8 upon complexation and reveal its segmental flexibility.

Two molecular dynamics simulations were carried out for the antibody Fab NC6.8, both with and without the guanidinium sweetener ligand NC174, in order to assess the segmental flexibility as well as the conformational changes upon ligand binding. Trajectory analyses of the simulation of the uncomplexed Fab suggest low-amplitude motions of the Ig domains with respect to each other, most clearly reflected by a periodic alteration of the elbow angle within a range of 11 degrees. Upon insertion of the hapten into the binding site, the quaternary structure of the Fab exhibits considerable rearrangements: the elbow angle changes by almost 30 degrees, the light chain is elongated and the heavy chain becomes more flexed. Comparison with experiment reveals some interesting agreements with X-ray crystallographic results published previously.

Catalysis of dialanine formation by glycine in the salt-induced peptide formation reaction.

Mutual catalysis of amino acids in the salt-induced peptide formation (SIPF) reaction is demonstrated for the case of glycine/alanine. The presence of glycine enhances dialanine formation by a factor up to 50 and enables dialanine formation at much lower alanine concentrations. The actual amounts of glycine play an important role for this catalytic effect, the optimal glycine concentration is 1/8 of the alanine concentration. The mechanism appears to be based on the formation of the intermediate Gly-Ala-Ala tripeptide, connected to one coordination site of copper(II) ion, and subsequent hydrolysis to dialanine and glycine.

The effect of smectite composition on the catalysis of peptide bond formation.

Clay-catalyzed glycine and diglycine oligomerizations were performed as drying/wetting cycles at 80 degrees C. Two trioctahedral smectites (hectorite and saponite), three pure montmorillonites, a ferruginous smectite, an Fe(II)-rich smectite, and three smectites containing goethite admixture were used as catalysts. Highest peptide bond formation was found with trioctahedral smectites. About 7% of glycine was converted to diglycine and diketopiperazine on hectorite after 7 days. In the case of dioctahedral smectites, highest yields were achieved using clays with a negative-layer charge localized in the octahedral sheets (up to 2% of converted glycine after 7 days). The presence of Fe(II) in clay is reflected in a higher efficiency in catalyzing amino acid dimerization (about 3.5% of converted glycine after 7 days). The possible significance of the results for prebiotic chemistry is discussed.

Comparative molecular field analysis of haptens docked to the multispecific antibody IgE(Lb4)

Using comparative molecular field analysis (CoMFA), three-dimensional quantitative structure-activity relationships were developed for 27 haptens which bind to the monoclonal antibody IgE(Lb4). In order to obtain an alignment for these structurally very diverse antigens, the compounds were docked to a previously modeled receptor structure using the automated docking program AUTODOCK (Goodsell, D.S.; Olson, A.J. Proteins: Struct., Funct., Genet. 1990, 8, 195-202). Remarkably, this alignment method yielded highly consistent QSAR models, as indicated by the corresponding cross-validated r2 values (0.809 for a model with carbon as probe atom, 0.773 for a model with hydrogen as probe atom). Conventional alignment failed in providing a basis for self-consistent CoMFAs. Amino acids Tyr H 50, Tyr H 52, and Trp H 95 of the receptor appeared to be of crucial importance for binding of various antigens. These findings are consistent with earlier considerations of aromatic residues being responsible for the multispecificity of certain immunoglobulins.

Comparative docking studies on ligand binding to the multispecific antibodies IgE-La2 and IgE-Lb4.

A large comparative study is presented in which the binding of approximately 30 different ligands to two IgE antibodies (La2 and Lb4) is analyzed by means of an automated-docking procedure based on simulated annealing. The method is able to reproduce experimentally verified binding orientations, as shown by application to the Ig-AN02-hapten complex. The main address of the study is to investigate the concept of antibody multispecificity. Problems and usefulness of docking in this context are discussed. The results indicate reasons for multispecific binding properties and how they can be understood from the topology of the binding site. Though similar in general behaviour, the two antibodies show interesting differences in their binding characteristics. The binding sites of both antibodies are described and the main interacting residues revealed.

Prediction of IgE(Lb4)-ligand complex structures by automated docking.

A mouse monoclonal anti-2,4,6-trinitrophenyl IgE (clone Lb4) was screened with a random set of over 2000 compounds, and several ligands were found to bind with affinities comparable to that of the immunizing hapten (KD in the microM range). An automated docking algorithm was used for the prediction of complex structures formed by 2,4-dinitrophenyl (DNP) and non-DNP ligands in the fragment variable region of IgE(Lb4). All ligands were found to dock in an L-shaped cavity of 15 x 16 x 10 A, surrounded by complementary-determining regions L1, L3, H2 and H3. The ligands were found to occupy the same binding site in different orientations. For rigid ligands the most stable orientation could be predicted with high probability, based on the calculated energy of binding and the occurrence frequencies of identical complexes obtained by repeated simulations. The localization of a flexible ligand (cycrimine-R) was more ambiguous, but it still docked in the same site. The results support a model for heteroligating antibody (Ab) binding sites, where different ligands utilize the total set of available contacts in different combinations. It is suggested that although pseudoenergies calculated by the docking algorithm do not correlate with experimentally measured binding energies, the screening-and-docking procedure can be useful for the mapping of Ab and other receptor binding sites ligating small molecules.

Heteroligation of a mouse monoclonal IgE antibody (La2) with small molecules, analysed by computer-aided automated docking.

A mouse monoclonal anti-TNP IgE antibody (IgE-La2) was screened by a competitive-binding ELISA with a random pool of over 2000 small molecules, mostly drugs, drug derivatives and metabolites. Thirteen of these (naproxene, beta-carboxy-chi-naphthol, oxolinic acid, hymecromone, 8-aminoquinoline, beta-naphthylamine, chi-nitrilo-cinnamic acid, 1,5-diaminonaphthaline, prolonium iodide, diaspirin, 3,4,5-trimethoxy-cinnamic acid, cycrimine, hemimellitic acid) were found to bind as strongly, or stronger, to the antibody as the immunizing hapten. We have used a Monte Carlo search technique for simulated docking of the DNP and non-DNP ligands to a model of the Fv region of IgE(La2). The validity of structural predictions made by the AutoDock program were tested on IgG(ANO2), the three-dimensional structure of which had been obtained previously by X-ray crystallography and 2D-NMR. The rms differences between the experimentally determined and auto-docked complexes in the energetically most favored binding modes were 0.31-0.44 A. Evaluation of structures of IgE(La2)-ligand complexes [including 2,4-dinitrophenol (DNP), 16 DNP amino acids, and the 13 non-DNP ligands listed above] obtained by computer-aided automated docking, suggested the existence of two subsites within an approximately 12 x 18 A2 groove extending between the H and L CDRs. Some of the ligands (DNP-Glu, 8-aminoquinoline, prolonium-I, beta-naphthylamine) were found to bind exclusively to subsite 1, others (DNP-Ala, chi-nitrilo-cinnamic acid, hemimellitic acid, beta-carboxy-chi-naphthol) to subsite 2. The majority of DNP amino acids and other ligands (oxolinic acid, 3,4,5-trimethoxy-cinnamic acid, diaspirin, [R]-cycrimine) were found to occupy an overlapping area including subsites 1 and 2, while some of the compounds (DNP-Asn, DNP-Pro, hymecromone, 1,5-naphthylenediamine) were predicted to interact with either of these subsites with comparable probabilities. When all of the docked La2-ligand complexes were taken into account, five tyrosine residues (H33, L32, L91, L92, L96) were found to provide the majority (53.4%) of all observed contact points. Thus, a multitude of interactions with aromatic residues, and a combinatorial type of interaction within the binding region, seem to be the major factors to explain the mechanism of heteroligation by IgE(La2).