Igor Tvaroška

Carbon-proton coupling constants in the conformational analysis of sugar molecules.

Publisher Summary This chapter examines the carbon–proton coupling constants involved in the conformational analysis of sugar molecules. The indirect spin–spin coupling is independent of molecular rotation. The coupling mechanism is known to involve the electron spins of the bonding electrons and is the result of a weak electron polarization. A proton signal has two 13C satellites, each having an intensity that is 0.5% of that of the corresponding total proton resonance and located symmetrically around that resonance. In addition to providing information about 1H ( 1H couplings) these satellites allow the determination of the 1JC,H couplings with good resolution. With the advent of inverse detection and the corresponding gain in sensitivity, newer schemes for the measurement of long-range coupling constants have been proposed. They are based on the heteronuclear multiple-bond correlation derivative or variants of the heteronuclear multiple-quantum coherence experiment, and also on the heteronuclear single-quantum coherence experiment. The Hamiltonian gives the total electronic kinetic energy and the magnetic interactions between electron orbital motions and nuclear magnetic moments. It is found that two-bond carbon–proton coupling constants are much smaller than one-bond couplings and their signs can be negative or positive.

Quantum mechanical and NMR spectroscopy studies on the conformations of the hydroxymethyl and methoxymethyl groups in aldohexosides.

The potential energy surfaces of the hydroxymethyl and methoxymethyl groups in methyl hexopyranosides have been extensively studied, employing quantum mechanical calculations and high resolution NMR data. The structure and energy of the C-5-C-6 rotamers were calculated at the B3LYP level of the density functional theory (DFT). For all, geometry optimizations were carried out for 264 conformers of 16 methyl D-gluco- and methyl D-galactopyranoside derivatives 1-16 at the B3LYP/6-31G** level. For all calculated minima, single-point calculations were performed at the B3LYP/6-311++G** level. Solvent effects were considered using a self-consistent reaction field method. Values of the vicinal coupling constants 3J(H-5-H-6R), 3J(H-5-H-6S), 3J(C-4-H-6R), and 3J(C-4-H-6S) for methyl D-glucopyranosides, methyl D-galactopyranosides and their 6-O-methyl derivatives 9-16 were measured in two solvents, methanol and water. The calculated gg, gt, and tg rotamer populations of the hydroxymethyl and methoxymethyl groups in 9-16 agreed well with experimental data. The results clearly showed that the population of gg, gt, and tg rotamers is sensitive to solvent effects. It was concluded that the preference of rotamers in 1-16 is due to the hydrogen bonding and solvent effects.

Modelling of β-d-glucopyranose ring distortion in different force fields: a metadynamics study

Modelling of carbohydrate conformations is a challenging task for force field developers. Three carbohydrate force fields, namely GLYCAM06, GROMOS 45a4 and OPLS were evaluated. Free energies of different ring conformations of beta-D-glucopyranose were calculated using metadynamics in vacuum as well as in explicitly modelled water. All three force fields model the (4)C(1) conformation as the most stable by at least 6kJ/mol, as compared to other conformations. Interconversion from the (4)C(1) to any other conformation is associated with a barrier of no lower than 26kJ/mol. The free energy surface calculated in the GLYCAM06 force field is in remarkably good agreement with the recent Car-Parrinello metadynamics study. The effect of a water environment is relatively low and analogous in all tested force fields. Namely, the presence of water stabilizes the upper-left ((3,O)B) versus bottom-right (B(3,O)) area of Stoddards plot, relative to the situation in vacuum. Comparison of free and potential surfaces is also provided for vacuum calculations.

Hydration of α-maltose and amylose: molecular modelling and thermodynamics study

Abstract Hydration of α-maltose and amylose were investigated using molecular modelling and thermodynamics methods. The structure and energy of hydration of three low-energy conformers of α-maltose were determined by the MM3 molecular mechanics method. The hydration structure was found to be sensitive to the conformation of α-maltose and hydration numbers 10 or 11 were estimated for the different conformers. Differential scanning calorimetry and thermogravimetric analysis were used to determine the number of water molecules specifically bonded (non-freezing water) to amylose and different samples of α-maltose. Due to high crystallinity of α-maltose samples, the observed non-freezing water content was lower than predicted by molecular modelling. In contrast, the experimental number of non-freezing molecules of water per d -glucopyranose residue for amorphous amylose (nh = 3.8) is in good accordance with the value of 3.8 extracted from our calculations.

Binding of β-amyloid (1-42) peptide to negatively charged phospholipid membranes in the liquid-ordered state: modeling and experimental studies.

To explore the initial stages of amyloid β peptide (Aβ42) deposition on membranes, we have studied the interaction of Aβ42 in the monomeric form with lipid monolayers and with bilayers in either the liquid-disordered or the liquid-ordered (L(o)) state, containing negatively charged phospholipids. Molecular dynamics (MD) simulations of the system have been performed, as well as experimental measurements. For bilayers in the L(o) state, in the absence of the negatively charged lipids, interaction is weak and it cannot be detected by isothermal calorimetry. However, in the presence of phosphatidic acid, or of cardiolipin, interaction is detected by different methods and in all cases interaction is strongest with lower (2.5-5 mol%) than higher (10-20 mol%) proportions of negatively charged phospholipids. Liquid-disordered bilayers consistently allowed a higher Aβ42 binding than L(o) ones. Thioflavin T assays and infrared spectroscopy confirmed a higher proportion of β-sheet formation under conditions when higher peptide binding was measured. The experimental results were supported by MD simulations. We used 100 ns MD to examine interactions between Aβ42 and three different 512 lipid bilayers consisting of palmitoylsphingomyelin, dimyristoyl phosphatidic acid, and cholesterol in three different proportions. MD pictures are different for the low- and high-charge bilayers, in the former case the peptide is bound through many contact points to the bilayer, whereas for the bilayer containing 20 mol% anionic phospholipid only a small fragment of the peptide appears to be bound. The MD results indicate that the binding and fibril formation on the membrane surface depends on the composition of the bilayer, and is the result of a subtle balance of many inter- and intramolecular interactions between the Aβ42 and membrane.

The conformational properties of the glycosidic linkage

Abstract Stereochemical properties of the glycosidic linkage have been studied by the quantum-chemical PCILO method, using 2-methoxytetrahydropyran as a model. Calculations of the two-dimensional, conformational (Φ, Ψ) maps showed that the rotation around the C-1O-1 bond is more hindered than that around the O-1C-6 bond, and that there are differences in the shape of the energy curve for the axial and equatorial forms of 2-methoxytetrahydropyran. The observed population of the five stable conformers at equilibrium (GG:GT:TG 1 :TG 2 :TT = 70.8:6.0:19.9:2.0:1.3) is consistent with the prediction of the anomeric and exo-anomeric effects. The calculated abundance (76.8%) of the axial form of 2-methoxytetrahydropyran is comparable with experimental results (77–80%) obtained by n.m.r. measurements in non-polar solvents. The energies found for individual conformers made it possible to calculate the magnitude of the anomeric effect (3 kJ/mol) and to determine, for the first time, the values of the exo-anomeric effect for axial (6 kJ/mol) and equatorial 2-methoxytetrahydropyran (7 kJ/mol). The calculated variations of the geometry arising from rotation around the C-1O-1 bond are consistent with results obtained by statistical analysis of experimental data for α- and β-glycosides. The results obtained, indicating that the energy, geometry, and electronic structure of glycosides are largely affected by the conformation of the acetal segment, are discussed from the point of view of conformational analysis of oligo- and poly-saccharides.

Hybrid Quantum Mechanical/Molecular Mechanical Investigation of the β-1,4-Galactosyltransferase-I Mechanism

The enzyme beta-1,4-galactosyltransferase-1 (beta4Gal-T1) catalyzes the transfer of a galactose residue from UDP-Gal to the C4-hydroxyl group of N-acetylglucosamine. The catalytic mechanism of beta4Gal-T1 was investigated using the hybrid quantum mechanical/molecular mechanical (QM/MM) method, with the QM portion containing 253 atoms treated with density functional theory (DFT) at the BP/DZP and BP/TZ2P levels. The remaining parts of the beta4Gal-T1 complex, 4527 atoms in all, were modeled using the AMBER molecular force field. A theoretical model of the Michaelis complex was built using the X-ray structure of beta4Gal-T1 in a complex with the donor or acceptor substrate, respectively. The hybrid QM(DFT)/MM calculations identified an S(N)2-type transition state for the nucleophilic attack of the O4(a) oxygen on the anomeric carbon C1 and the breaking of the C1-O1 glycosidic linkage. The activation barrier found for this process is 15 kcal/mol. In the transition state (TS) model, the sugar donor is partially cleaved from pyrophosphate, while nucleophilic oxygen O4(a) remains protonated with a low barrier hydrogen bond to the catalytic base D318. The structure of TS is characterized by the O4(a)-C1 and C1-O1 distances of 2.703 and 2.092 A, respectively. When the obtained reaction sequence was used, the nature of the captured intermediate resembling the transition state structure (PDB/2FYD) was elucidated. This modeling QM/MM study has provided detailed insight into the mechanism of the Gal transfer catalyzed by beta4Gal-T1 and has supplied further evidence for a concerted S(N)2-type displacement mechanism employed by inverting glycosyltransferases.

Dependence on saccharide conformation of the one-bond and three-bond carbonproton coupling constants☆☆☆

Abstract A theoretical study is presented of the dependence on conformation of 1 J C,H and 3 J C,H values in model compounds related to glycosides. Calculated J values for dimethoxymethane and 2-methoxytetrahydropyran are based on the FPT formulation in the semiempirical INDO method. The configuration at the anomeric carbon affects the 1 J C;H value, and the 1 J C,H and 3 J C,H values vary characteristically with dihedral angle about the carbonoxygen bond. The agreement of the calculated and experimental values is satisfactory, especially for the 3 J C,H values.

Comparative DFT study on the α-glycosidic bond in reactive species of galactosyl diphosphates

Correct prediction of the structure and energetics along the reaction pathway of the formation or dissociation of the glycosidic bond in sugar phosphates is crucial for the understanding of catalytic mechanism and for the determination of transition state structures of sugar-phosphate processing enzymes. The performance of seven density functional theory (DFT) methods (BLYP, B3LYP, MPW1PW91, MPW1K, MPWB1K, M05 and M05-2X) and two wave function methods (HF and MP2) was tested using four structural models with the activated sugar-phosphate α-glycosidic linkage. The models were chosen based on the crystal structure of the retaining glycosyltransferase LgtC complex with methyl α-d-galactopyranose diphosphate and its 2-fluoro derivative. Results of the MP2 method were used as a benchmark for the other methods. Two structural trends were observed in the calculations: predicted length of the activated C1-O1 glycosidic bond of 1.49–1.63 Å was significantly larger than values of a standard C1-O1 glycosidic bond in crystal structures of carbohydrates (1.39–1.48 Å), and the calculated value depended on the DFT method used. The MPW1K, M05 and M05-2X functionals provided results in closest agreement with those from the MP2 method, the difference being less than 0.02 Å in the calculated glycosidic bond lengths. On the contrary, the BLYP and B3LYP functionals failed to predict sugar diphosphate in the (-sc) conformation as a stable structure. Instead, the only stationary points localized along the C1-O1 dissociation coordinate were oxocarbenium ions at the distance of approximately 2.8 Å. The M05-2X, MPW1K and MPWB1K functionals gave the most reasonable prediction of the thermochemical kinetic parameters, where the formation of the oxocarbenium ion has a slightly endothermic character (0.4–1.7 kJ mol−1) with an activation barrier of 7.9–9.2 kJ mol−1.

Angular dependence of vicinal carbon-proton coupling constants for conformational studies of the hydroxymethyl group in carbohydrates☆

Abstract A theoretical study is presented of the dependence on the hydroxymethyl group conformation of vicinal carbon-proton coupling constant 3 J C,H in a series of 16 hexopyranoses. Calculated 3 J C,H values for both anomers of d -glucopyranose (1), d -mannopyranose (2), d -allopyranose (3), d -altropyranose (4), d -galactopyranose (5), d -talopyranose (6), d -gulopyranose (7), and d -idopyranose (8) are based on the FTP formulation in the semi-empirical approximation of INDO. The dependence of the coupling constants on the dihedral angle ω C between the coupling carbon atom C-4 and protons H-6 is represented by a trigonometric function of the form 3 J C,H = 5.8cos 2 ω C − 1.6cos ω C + 0.28sin2 ω C − 0.02sin ω C + 0.52. It was found that the configuration at the anomeric and C-4 carbon atoms does not show any significant influence on 3 J C,H values. Agreement of calculated and experimental values available for mono- and oligo-saccharides is satisfactory. Based on these results, it is concluded that proposed equation for 3 J C,H values can be used as a tool for estimation of the conformational properties of the hydroxymethyl group in monosaccharides and of (1 → 6)- linked oligosaccharides in solution.