Modeling of arabinofuranose and arabinan,II. Nmr and Conformational analysis of arabinobiose and arabinan

The disaccharide arabinobiose (5-O -α-L -arabinofuranosyl-α-L -arabinofuranose) constitutes the basic repeating structures found in such polysaccharides as arabinan or in the side chains of the hairy regions of pectins. The conformational behavior of aqueous arabinobiose has been investigated by high resolution nmr and computerized molecular modeling. The complete conformational analysis of the, disaccharide has been achieved with the MM3 molecular mechanics methods using the flexible residue method. In this study, both the puckering of the arabinofuranose, rings and the orientations about the glycosidic torsion angles ?, ψ, and ω; were considered. Some insights into conformational transitions were obtained through molecular dynamics simulation using the CHARMM force field. In parallel, transient nuclear Overhauser effects at 400.13 MHz and long-range vicinal homonuclear and heteronuclear coupling constants have been measured. The theoretical nmr data were calculated taking into account all accessible conformations and using averaging methods for both slow and fast internal motions models. The data do not support a single conformational model, and only conformational averaging yields the excellent agreement between the observed and simulated parameters. Within the potential energy surfaces computed for the disaccharide, several low energy conformers can be identified. When these conformations are extrapolated to regular polysaccharide structures, they generate chains of arabinan displaying right- and left-handed chirality and a wide range of repeating units per turn of helix. © 1994 John Wiley & Sons, Inc.     

Conformational-energy calculations for oligosaccharides: a comparison of methods and a strategy of calculation

A theoretical conformational analysis of dimethoxymethane, 2-methoxytetrahydropyran, cellobiose, and maltose has been performed. The validity of several commonly used classical approaches to conformational energy, assuming non-bonded interactions, torsional terms, and the exo-anomeric contribution, and the MM2CARB method (a modified version of the MM2 force-field program using the Jeffrey-Taylor parameters) was tested against available experimental data or previous quantum-chemical calculations. The MM2CARB method correctly reproduces the energies and the variations in bond lengths and bond angles for conformers of dimethoxymethane and 2-methoxytetrahydropyran. Prediction of the observed conformers with simple potential functions appears to be less satisfactory. In particular, calculations that take into account non-bonded interactions and the exo-anomeric contribution based on dimethoxymethane give predicted energy differences that are 2–3 times higher than the experimental values. The general shapes of the (Φ, Ψ) potential-energy surfaces for cellobiose and maltose provided by potential-function calculations suggest the presence of several minima whose energies depend, to a great extent, on the choice of molecular geometry. The MM2CARB-calculated structures of seven cellobiose and five maltose conformers demonstrate clearly the variation of disaccharide geometry with change of conformation around the glycosidic linkage. The relative energies calculated by simple methods differ from MM2CARB energies and indicate that the simple potential-functions methods give only a qualitative estimate of oligosaccharide conformers. Based on these results, we propose a general strategy and two different approaches for the investigation of conformational properties of oligosaccharides.     

Three-dimensional features of the bacterial polysaccharide (1 → 4)-β-D-glucuronan: A molecular modeling study

The mutant strain M5N1 CS of Rhizobium meliloti produces, in a Rhizobium complete medium supplemented with fructose and sucrose, a partially acetylated homopolymer of D -glucuronic acid residues linked β-(1 → 4). This polysaccharide forms thermoreversible gels with monovalent salts and thermally stable gels with divalent salts. In order to define the different levels of structural characterization, modeling simulations were performed for both the regular (1 → 4)-β-D -glucuronan and the acetylated derivatives. This required the evaluation of the accessible conformational space for the 16 disaccharides. Detailed conformational analysis was accomplished using the flexible residue of the MM3 molecular mechanics procedure and the results were used to access the configurational statistics of representative polysaccharide chains. Within the potential energy surfaces calculated for each disaccharide, several low energy conformers can be identified. When these conformations are extrapolated to regular polysaccharide structures, they generate polymers with right- and left-handed chirality along with a 2-fold axis. This later arrangement (n = 2, h = 5.16 Å) closely corresponds to that derived from a fiber x-ray diffraction investigation. The insertion of acetyl groups induces changes in the helical features of the polymer. As for the simulation of the configurational properties of (1 → 4)-β-D -glucuronan, an extended disordered chain having a persistence length of 105 Å (corresponding to 22 monomers) is predicted. This agrees with previous conclusions derived from solution study. The inclusion of varying amounts of acetyl groups only slightly perturbs the calculated persistence length. © 1998 John Wiley & Sons, Inc. Biopoly 45: 165–175, 1998     

Conformational analysis of the anomeric forms of kojibiose, nigerose, and maltose using MM3

Energy surfaces were computed for relative orientations of the relaxed pyranosyl rings of the two anomeric forms of kojibiose, nigerose, and maltose, the (1 → 2)-, (1 → 3)-- and (1 → 4)--linked -glucosyl disaccharides, respectively. Twenty-four combinations of starting conformations of the rotatable side-groups were considered for each disaccharide. Optimized structures were calculated using MM3 on a 20° grid spacing of the torsional angles about the glycosidic bonds. The energy surfaces of the six disaccharides were similar in many respects but differed in detail within the low-energy regions. The maps also illustrate the importance of the exo-anomeric effect and linkage type in determining the conformational flexibility of disaccharides. Torsional conformations of known crystal structures of maltosyl-containing molecules lie in a lower MM3 energy range than previously reported.     

Studies on the molecular recognition of synthetic methyl beta-lactoside analogs by ricin, a cytotoxic plant lectin

The binding of methyl beta-lactoside and of all possible monodeoxy derivatives of methyl beta-lactoside to the galactose-specific highly cytotoxin lectin ricin, has been investigated. The distribution of low-energy conformers of the disaccharide structures has been first determined using molecular-mechanics calculations and high-resolution NMR spectroscopy. The nuclear Overhauser enhancements and specific deshieldings observed are in agreement with a similar distribution of low-energy conformers for all studied compounds which may be described by a major conformer defined by phi (H1'-C1'-O1'-C4) and psi (C1'-O1'-C4-H4) torsion angles of 49 degrees and 5 degrees, respectively, with contribution of conformers with angles phi/psi 24 degrees/-59 degrees, 22 degrees/-32 degrees and 6 degrees/-44 degrees. Assuming that the disaccharides bind to the lectin in these preferred conformations, the apparent dissociation constants for the ricin-disaccharide complexes have been interpreted in terms of specific polar and nonpolar interactions. In agreement with X-ray data, the hydroxyl groups at positions 3, 4 and 6 of the beta-D-galactopyranose moiety appear as key polar groups in the interaction with ricin. These results are in contrast to previous results which have established that position 6 is not involved in lectin binding. An important nonpolar interaction involving position 3 of the beta-D-glucopyranose moiety, seems to be operative. The distribution of low-energy conformers of these disaccharide structures permits this interaction to take place with the hydroxyl group at this position intramolecularly bonded, thus rendering this region of the molecule more lipophylic in character for acceptance into nonpolar regions of the combining site.     

NMR and conformational studies of linear and cyclic oligo-(1→6)-β-d-glucosamines

The conformational behavior of a series of linear and cyclic oligo-(1→6)-β-d-glucosamines and their N-acetylated derivatives, which are related to fragments of natural poly-N-acetylglucosamine, was studied by theoretical molecular modeling and experimental determination of transglycosidic vicinal coupling constants ³J_C,H and ³J_H,H. Molecular dynamics simulations were performed under several types of conditions varying in the consideration of ionization of amino groups, solvent effect, and temperature. Neural network clustering and asphericity calculations were performed on the basis of molecular dynamics data. It was shown that disaccharide fragments in the studied linear oligosaccharides were not rigid, and tended to have several conformers, thus determining the overall twisted shape with helical elements. In addition, it was found that the behavior of C5–C6 bond depended significantly upon the simulation conditions. The cyclic di-, tri-, and tetrasaccharides mostly had symmetrical ring-shaped conformations. The larger cycles tended to adopt more complicated shapes, and the conformational behavior of their disaccharide fragments was close to that in the linear oligosaccharides.     

A conformational analysis of biologically active RGD-containing cyclopentapeptides

The theoretical conformational analysis of the biologically active RGD-containing pentapeptide cyclo(-Arg-Gly-Asp-Phe-DVal-), an inhibitor of laminin P1 interaction with its receptor, was performed. The space of permissible torsional angles of the backbone of the molecule was studied by the Monte Carlo method. From the large number of predicted low-energy conformers with various packings of the cyclic moiety of this pentapeptide, only those were selected that corresponded to stable structures of the model linear tripeptide Ac-Ala-Gly-Asp-NHMe. This peptide simulated the spatial possibilities of the backbones of RGD-containing fragments of laminin, vitronectin, and fibronectin. We selected several dozen structures that may be potential biologically active conformers, but only a few of them were capable of forming stable intramolecular hydrogen bonds. We assumed that a biologically active conformer of cyclo(-Arg-Gly-Asp-Phe-DVal-) can be present in significant amounts in an equilibrium mixture in solution along with other conformers without necessarily dominating among them.     

Conformational analysis of octa- and tetrabromo tetraphenylporphyrins and their Ni(II) and Tb(III) complexes

Molecular mechanics (MM) calculations were used to analyze the puckering of metalloporphyrins as a function of metal ion size and the position of substituents on the porphyrin periphery, on a three series of octa- and tetrabromo tetraphenylporphyrins: without metal, and with Ni(II), and Tb(III) as representative small and large metal ions, respectively. Molecular energy optimization calculations were carried out using the Consistent Force Field (CFF) program, with the parameters developed previously and new parameters for bromine atom. Normal-coordinate structural decomposition (NSD) analysis was performed on the equilibrium structures obtained by MM calculations. The conformers are also stereochemically characterized, compared with available X-ray structures and with the conformers obtained in our previous MM study using chloro instead of bromo beta-pyrrole substituents.     

Solvent effect on the stability of mannobiose conformers

The conformational equilibria of seven methyl β-D -mannobioside conformers have been studied theoretically in five solvents. The structure of each individual conformer has been refined by the PCILO quantum-chemical method from the seven distinct low-energy regions determined from (Φ, Ψ) maps calculated by a potential function method. The stability of the conformers in dilute solution has been evaluated by using a method that consists of electrostatic, dispersion, and cavity terms. The calculated abundance of conformers depends on the solvent and results indicate that the preponderant conformer in the solution may not be the one adopted by mannobiose in the crystalline form. Based on the determined abundance of conformers, thermodynamically averaged nmr parameters, dipole moment, and linkage rotation have been calculated. The solvation behavior of methyl β-D -mannobioside is compared to those previously estimated for cellobiose and maltose.     

Carcinogen-base stacking and base-base stacking in dCpdG modified by (+) and (-) anti-BPDE

The low-energy conformations accessible to dCpdG modified at guanine N² via trans epoxide opening by (+) and (?) 7β,8α-dihydroxy-9α,10α-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene (anti-BPDE) have been delineated by minimized semiempirical potential-energy calculations with all torsion angles flexible. Nearly 4000 trials were made, representing a fairly thorough investigation of the conformation space of the adducts. Carcinogen–base stacked states and base–base stacked conformers were found in the low-energy regions of both enantiomers. Many ω′, ω, ψ domains accommodate the two types of conformations, with B-like backbones among the most preferred states in each case. The conformational differences between the two enantiomers on the dimer level reside in subtle distinctions in orientation of the carcinogen–base linkage.     

Relaxed-residue conformational mapping of the three linkage bonds of isomaltose and gentiobiose with MM3(92)

Isoenergy surfaces were calculated for the α- and β-anomers of isomaltose and gentiobiose, based on 46,656 conformers for each disaccharide. Low-energy regions exist for each of the three staggered positions about the C-5′ ? C-6′ bonds, and known crystal structures lie in two of these regions. As expected, the molecular partition function showed greater flexibility for these three-bond-linked disaccharides than for comparable two-bond-linked structures. A model miniature crystal of gentiobiose accounts for most of the remaining structural differences between the modeled isolated molecule and the crystal structure. Based on models of isolated molecules of isomaltose and gentiobiose, the predicted Boltzmann-weighted nmr coupling constants were satisfactory, as were predicted optical rotations for gentiobiose. © 1994 John Wiley & Sons, Inc.     

Increased glycosidic bond stabilities in 4-C-hydroxymethyl linked disaccharides

Three new hydroxymethyl-linked non-natural disaccharide analogues, containing an additional methylene group in between the glycosidic linkage, were synthesized by utilizing 4-C-hydroxymethyl-α-d-glucopyranoside as the glycosyl donor. A kinetic study was undertaken to assess the hydrolytic stabilities of these new disaccharide analogues toward acid-catalyzed hydrolysis, at 60 °C and 70 °C. The studies showed that the disaccharide analogues were stable, by an order of magnitude, than naturally-occurring disaccharides, such as, cellobiose, lactose, and maltose. The first order rate constants were lower than that of methyl glycosides and the trend of hydrolysis rate constants followed that of naturally-occurring disaccharides. α-Anomer showed faster hydrolysis than the β-anomer and the presence of axial hydroxyl group also led to faster hydrolysis among the disaccharide analogues. Energy minimized structures, derived through molecular modeling, showed that dihedral angles around the glycosidic bond in disaccharide analogues were nearly similar to that of naturally-occurring disaccharides.     

Solution conformation of sialosylcerebroside (GM4) and its NeuAc(alpha 2----3)Gal beta sugar component

The solution conformations of GM4 ganglioside [NeuAc(alpha 2----3)Gal (beta 1----1)Cer] in (2H3C)2SO and its component disaccharide in 2H2O were investigated with the aid of 1H-nuclear magnetic resonance spectroscopy (nuclear Overhauser effect, analysis of coupling constants) and by energy-minimum calculations. The existence of three low-energy conformers obtained by theoretical calculations was supported by experimental findings in the case of GM4, whereas the disaccharide appears to exist as a mixture of two conformers.     

Influence of local interactions on protein structure. III. Conformational energy studies of N-acety-N′-methylamides of Gly-X and X-Gly dipeptides

Conformational energy calculations using an empirical conformational energy program for peptides (ECEPP) were carried out on 20 N-acetyl- N′-methylamides of Gly-X and X-Gly depeptides, where X = Ala, Asn, Asp, Gly, Phe, Ser, Thr, Tyr, Val, and Pro, and also of Leu-Gly. Each depeptde was found to have 25 or more low-energy minima, except Gly-Thr, which had only 11 low-energy minima because of the stable side chian-backbone hydrogen present in all low-energy conformation. As a group, the stble chain-backbone hydrogen bonds present in all low-energy conformations. As a group, the Gly-containing dipeptides were calculated in all low-energy prpensity for formation of bends than the Ala-containing depeptides. The X- Gly dipeptides were calculated to favor bends more than the Gly-X dipeptides, primarlly because of the high stability of the type II bend in X-Gly dipeptides. These results are in agreement with obseved occurrences of bends in the x-ray structures of globular proteins. The calculated conformation properties were found to be in good agreement with experimental results.     

Conformational energy calculations of enzyme-substrate complexes of lysozyme. I. Energy minimization of monosaccharide and oligosaccharide inhibitors and substrates of lysozyme

Conformational energy calculations were performed on monosaccharide and oligosaccharide inhibitors and substrates of lysozyme to examine the preferred conformations of these molecules. A grid-search method was used to locate all of the low-energy conformational regions for N-acetyl-β-D -glycosamine (NAG), and energy minimization was then carried out in each of these regions. Three stable positions for the N-acetyl group have ben located, in two of which the plane of the amide unit is normal to the mean plane of the pyranosyl ring. Nine local energy minima were located for the —CH₂OH group. The positions of the two vicinal cis —OH groups are determined predominantly by interactions with either the —CH₂OH or the N-acetyl group. The most stable conformations of β-N-acetylmuramic acid (NAM) were determined from the study of the low-energy conformations of NAG. In the two stable orientations for the D -lactic acid side chain, the O—C—C′ plane (C′ being the carbon atom of the terminal carboxyl group) was found to be normal to the mean plane of the pyranosyl ring. The low-energy positions for the COOH group of NAM are determined mainly by interactions with neighboring groups. The conformational preferences of the α-anomers of NAG and NAM were also explored. The calculated conformation of the N-acetyl group for α-NAG was quite close to that determined by X-ray analysis. Two of the three lowest energy conformations of α-NAM are similar to the corresponding conformations of the β-anomer. A third low-energy structure, which has a hydrogen bond from the NH of the N-acetyl group to the C?O of the lactic acid group, corresponds very closely to the X-ray structure of this molecule. The preferred conformations of the disaccharides NAG–NAG, NAM–NAG and NAG–NAM were also investigated. Two preferred orientations of the reducing pyranosyl ring relative to the nonreducing ring were found for all of these disaccharides, both of which are close to the extended conformation. In one of these conformations, a hydrogen bond can form between the OH group attached to C₃ of the reducing sugar and the ring oxygen of the preceding residue. Each conformation can be stabilized further by a hydrogen bond between the CH₂OH (donor) of residue i + 1 and the C?O of residue i (acceptor). The interactions that determine conformations for all oligosaccharides containing both NAG and NAM are shown to be exclusively intraresidue and nearest neighbor interactions, so that it is possible to predict all stable conformations of oligosaccharides containing NAG and NAM in any sequence.     

Comparison of different force fields for the study of disaccharides

Eighteen empirical force fields and the semi-empirical quantum method PM3CARB-1 were compared for studying β-cellobiose, α-maltose, and α-galabiose [α-d-Galp-(1→4)-α-d-Galp]. For each disaccharide, the energies of 54 conformers with differing hydroxymethyl, hydroxyl, and glycosidic linkage orientations were minimized by the different methods, some at two dielectric constants. By comparing these results and the available crystal structure data and/or higher level density functional theory results, it was concluded that the newer parameterizations for force fields (GROMOS, GLYCAM06, OPLS-2005 and CSFF) give results that are reasonably similar to each other, whereas the older parameterizations for Amber, charmm or OPLS were more divergent. However, MM3, an older force field, gave energy and geometry values comparable to those of the newer parameterizations, but with less sensitivity to dielectric constant values. These systems worked better than MM2 variants, which were still acceptable. PM3CARB-1 also gave adequate results in terms of linkage and exocyclic torsion angles. GROMOS, GLYCAM06, and MM3 appear to be the best choices, closely followed by MM4, CSFF, and OPLS-2005. With GLYCAM06 and to a lesser extent, CSFF, and OPLS-2005, a number of the conformers that were stable with MM3 changed to other forms.     

Disaccharide conformational maps: how adiabatic is an adiabatic map?

The 'adiabatic' (phi, psi) potential-energy surface of the disaccharide alpha-D-galactopyranosyl-(1-->3)-beta-D-galactopyranose was obtained by several established methods, using the MM3 molecular mechanics force field. The constrained minimizations throughout the whole grid were carried out using sharply different dielectric constants. The attainment of the 'true' adiabatic map is very difficult due to the 'multiple minimum problem', originating in the large number of exocyclic pendant groups present in a disaccharide. However, these results suggest that at low dielectric constants, the usual approach starting with conformers carrying cooperative hydrogen bonds results in a good approximation to the true adiabatic map, while at high dielectric constants this approach fails due to the damping of electrostatic and hydrogen-bonding interactions.     

Crystal Structure of the GalNAc/Gal-Specific Agglutinin from the Phytopathogenic Ascomycete Sclerotinia sclerotiorum Reveals Novel Adaptation of a β-Trefoil Domain

A lectin from the phytopathogenic ascomycete Sclerotinia sclerotiorum that shares only weak sequence similarity with characterized fungal lectins has recently been identified. S. sclerotiorum agglutinin (SSA) is a homodimeric protein consisting of two identical subunits of ∼ 17 kDa and displays specificity primarily towards Gal/GalNAc. Glycan array screening indicates that SSA readily interacts with Gal/GalNAc-bearing glycan chains. The crystal structures of SSA in the ligand-free form and in complex with the Gal-β1,3-GalNAc (T-antigen) disaccharide have been determined at 1.6 and 1.97 Å resolution, respectively. SSA adopts a β-trefoil domain as previously identified for other carbohydrate-binding proteins of the ricin B-like lectin superfamily and accommodates terminal non-reducing galactosyl and N-acetylgalactosaminyl glycans. Unlike other structurally related lectins, SSA contains a single carbohydrate-binding site at site α. SSA reveals a novel dimeric assembly markedly dissimilar to those described earlier for ricin-type lectins. The present structure exemplifies the adaptability of the β-trefoil domain in the evolution of fungal lectins.