Karim Mazeau

Overview on structural characterization of chitosan molecules in relation with their behavior in solution.

This paper concerns the new results obtained on the characterization of chitins and chitosans. Large series of samples was analyzed covering a wide range of water soluble and insoluble materials. The water soluble polymers were obtained by heterogeneous deacetylation and by homogeneous reacetylation. The calibration of IR spectrum was proposed and shown to be valid in all the range of DA. Application of 15 N and C solid state NMR was developed to be able to determine DA even in situ on insoluble natural materials. All the methods proposed give a very coherent set of results, The molecular weight distribution was established by GPC using cationic porous supports and the good solvent earlier proposed 0.3M AcOH/ 0.2M AcONa. The role of the distribution of acetyl groups along the chain is also discussed and analyzed by NMR ; it is demonstrated clearly the difference between homogeneously acetylated samples and heterogeneous samples coming from different routes of preparation. The dependence of Mark-Houwink parameters allowing to relate the intrinsic viscosity with the molecular weight is also briefly reported. In good concordance with experimental data, molecular modeling helps in understanding the role of the N-acetyl group content and distribution on the stiffness of the chains.

Conformational analysis of galactomannans: from oligomeric segments to polymeric chains

Abstract Conformational features of the glycosidic bond linking two mannosyl units of four different oligomeric fragments of galactomannans have been calculated by means of adiabatic mapping of the glycosidic Φ, Ψ torsion angles using the MM3 force field. These fragments differed in their substitution pattern. The aim of this study was to ascertain the role played by the galactosyl side groups on the conformational flexibility of the galactomannan chain backbone. Although the overall features of all the potential energy surfaces created appear similar, these maps show that the position of the lowest energy minimum conformer and the lower energy region change significantly if one or both mannosyl residues are substituted by a galactosyl side group. Thus, these groups lead to significant differences in the accessible conformational space, when compared with that of the mannobiose molecule. Predicted homonuclear and heteronuclear coupling constants averaged over each entire map also reflect the conformational differences. Computed maps were used to predict polymeric unperturbed dimensions, C ∞ , a , 〈 R 〉 and 〈 s 2 〉 1/2 of idealized galactomannan chains by Monte Carlo methods. For low values of Man:Gal ratios, chain extension appears to be strongly dependent on the degree of substitution. For 2:1 and 3:1 Man:Gal ratios, random, alternate and block patterns of substitution have been investigated. It has also been shown that the spatial extension of the polymer chains is dependent on the scheme of substitution. Such studies provide a unique insight into the dependence of these two factors on the stiffness and flexibility of different galactomannan chains.

Predicted Influence of N-Acetyl Group Content on the Conformational Extension of Chitin and Chitosan Chains

ABSTRACT Conformational analysis of chitosan molecules has been performed using the MM3(92) force field to investigate the role played by the acetamido groups on the stiffness of these chains. A high dielectric constant value was needed to model an aqueous environment and to reproduce the distribution of the N-acetyl glucosamine group orientation that is observed by NMR. Disaccharidic fragments, differently substituted at C2, were selected as models for chitin and chitosan chains. Their conformational space has been explored by means of adiabatic mapping of the glycosidic Φ,Ψ torsion angles. Although the overall features of all the potential energy surfaces created appear similar, the accessible conformational space of a glycosidic bond is affected by the nature of the substituent at C2 on the non-reducing residue of the disaccharide unit. This is illustrated by the differences in the calculated partition functions together with the predicted average homonuclear and heteronuclear coupling constants. Computed maps were used to predict polymeric unperturbed dimensions, characteristic ratio and persistence length of idealized chitin and chitosan chains, by Monte Carlo methods. Pure chitosan is predicted to be more coiled than pure chitin chains. At low N-acetyl group contents, chain extension appears to be dependent on the degree of substitution. Average chain dimensions increase monotonically for increases in content up to 60% of N-acetyl groups, but show no significant variation at higher contents. For molecules consisting of 50% amino and 50% N-acetylated residues, random, alternate and block patterns of substitution have been investigated. It has also been shown that the spatial extension of the polymer chains is dependent on the primary structure. Comparison with the literature experimental data is difficult because of the extreme diversity of the reported conformationally dependent values. However, such study provides a unique insight into the dependence of these two factors (degree of acetylation and distribution of acetyl groups) on the stiffness and flexibility of different chitin and chitosan chains.

Dynamics of cellulose-water interfaces: NMR spin-lattice relaxation times calculated from atomistic computer simulations.

Solid-state nuclear magnetic resonance (CP/MAS 13C NMR) spectroscopy has often been used to study cellulose structure, but some features of the cellulose NMR spectrum are not yet fully understood. One such feature is a doublet around 84 ppm, a signal that has been proposed to originate from C4 atoms at cellulose fibril surfaces. The two peaks yield different T1, differing by approximately a factor of 2 at 75 MHz. In this study, we calculate T1 from C4-H4 vector dynamics obtained from molecular dynamics computer simulations of cellulose I beta-water interfacial systems. Calculated and experimentally obtained T1 values for C4 atoms in surface chains fell within the same order of magnitude, 3-20 s. This means that the applied force field reproduces relevant surface dynamics for the cellulose-water interface sufficiently well. Furthermore, a difference in T1 of about a factor of 2 in the range of Larmor frequencies 25-150 MHz was found for C4 atoms in chains located on top of two different crystallographic planes, namely, (110) and (10). A previously proposed explanation that the C4 peak doublet could derive from surfaces parallel to different crystallographic planes is herewith strengthened by computationally obtained evidence. Another suggested basis for this difference is that the doublet originates from C4 atoms located in surface anhydro-glucose units with hydroxymethyl groups pointing either inward or outward. This was also tested within this study but was found to yield no difference in calculated T1.

A priori crystal structure prediction of native celluloses

The packing of beta-1,4-glucopyranose chains has been modeled to further elaborate the molecular structures of native cellulose microfibrils. A chain pairing procedure was implemented that evaluates the optimal interchain distance and energy for all possible settings of the two chains. Starting with a rigid model of an isolated chain, its interaction with a second chain was studied at various helix-axis translations and mutual rotational orientations while keeping the chains at van der Waals separation. For each setting, the sum of the van der Waals and hydrogen-bonding energy was calculated. No energy minimization was performed during the initial screening, but the energy and interchain distances were mapped to a three-dimensional grid, with evaluation of parallel settings of the cellulose chains. The emergence of several energy minima suggests that parallel chains of cellulose can be paired in a variety of stable orientations. A further analysis considered all possible parallel arrangements occurring between a cellulose chain pair and a further cellulose chain. Among all the low-energy three-chain models, only a few of them yield closely packed three-dimensional arrangements. From these, unit-cell dimensions as well as lattice symmetry were derived; interestingly two of them correspond closely to the observed allomorphs of crystalline native cellulose. The most favorable structural models were then optimized using a minicrystal procedure in conjunction with the MM3 force field. The two best crystal lattice predictions were for a triclinic (P(1)) and a monoclinic (P2(1)) arrangement with unit cell dimensions a = 0.63, b = 0.69, c = 1.036 nm, alpha = 113.0, beta = 121.1, gamma = 76.0 degrees, and a = 0.87, b = 0.75, c = 1.036 nm, gamma = 94.1 degrees, respectively. They correspond closely to the respective lattice symmetry and unit-cell dimensions that have been reported for cellulose Ialpha and cellulose Ibeta allomorphs. The suitability of the modeling protocol is endorsed by the agreement between the predicted and experimental unit-cell dimensions. The results provide pertinent information toward the construction of macromolecular models of microfibrils.

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.

Wetting the (110) and (100) Surfaces of Iβ Cellulose Studied by Molecular Dynamics

When observing the molecular details of the surfaces of cellulose microfibrils, one is struck by their heterogeneity, indicating that there are areas that are purely hydrophilic, whereas others have the hydrophobic character. Nature benefits from this ambivalence in many ways. For instance in plant cell walls, cellulose is able to interact not only with hydrophilic adducts such as hemicelluloses but also with the hydrophobic aromatic moieties of lignin. Other hydrophobic interactions occur when the aromatic part of a cellulase strongly binds to cellulose with the help of the so-called “cellulose-binding module”. In the case of the I cellulosesthe most abundant allomorph in higher plantssthe external morphology of the crystalline microfibrils corresponds mostly to the (110) and (11j0) surfaces, which are highly decorated by protruding hydroxyls. As a consequence, cellulose possesses a dominant hydrophilic character, which makes it incompatible with most synthetic polymers that are hydrophobic. In addition to these dominant hydrophilic surfaces, the (100) surface is believed to be much less represented. Since this surface exposes exclusively hydrophobic C-H moieties to the surrounding medium, cellulose also possesses the additional property to capture nonpolar solvents such as chloroform or cyclohexane. It is also this (100) surface that is responsible for the cellulases initial adsorption and further attack. The measurement of the contact angle of water drops deposited on a surface is the traditional method to determine the hydrophilic/hydophobic character of this surface. The test consists in measuring at the contact point the angle between the tangent drawn at the drop surface and that of the tangent of the supporting surface. In addition to the significance of this angle, the surface energy may also be derived from these measurements. Given the importance of the wettability of cellulosics, contact angles and their associated surface energies have been extensively measured for native and regenerated cellulose. The contact angle is a macroscopic parameter; its value gives an average of the surface characteristic but does not account for the chemical heterogeneity of the surface. The hysteresis effect often observed in dynamical wetting experiments partly accounts for this heterogeneity, but this phenomenon is difficult to interpret as hysteresis depends on many other factors. In an attempt to better characterize the surface anisotropy of cellulose, Yamane and his collaborators have recently demonstrated that the wetting ability of regenerated cellulose toward water could be lowered to some extent when the samples had been dipped into liquids of low dielectric constant. According to these authors, the ensuing increase in contact angle was likely due to a rearrangement of the hydrophilic surfaces, which consequently became more hydrophobic. The molecular mechanism of such rearrangement, if it can be proven, involves the rotation of some of the cellulose surface chains (and probably some of those in the crystal core as well) around their molecular axis. With the goal of ultimately verifying this debatable hypothesis, we have undertaken a preliminary study devised to mimic the wetting of cellulose surfaces by water, using atomistic molecular dynamics simulations. The question of the theoretical wetting limit of cellulose is addressed by considering separately the surfaces (110) and the (100) of the I allomorph.

The three-dimensional structure of the mega-oligosaccharide rhamnogalacturonan II monomer: a combined molecular modeling and NMR investigation.

In this study, we describe the first optimized molecular models of the mega-oligosaccharide rhamnogalaturonan II, that is found in the primary cell walls of all higher plants. The 750 MHz 1H NMR data previously reported and new heteronuclear correlation spectra (sensitivity-enhanced HSQC and HSQC-TOCSY) were first reassigned in light of the modifications in the primary structure. In turn, the experimental NMR data revealed the presence of an additional sugar, alpha-Araf (E-chain), and also the disaccharidic repeating unit of RG-I, another component of the pectic matrix. Due to a fuller picture of the primary structure of RG-II, a much more complete assignment of the NOE data has been achieved. A systematic computational study based on these NOEs lead us to a realistic three-dimensional description of the RG-II, in excellent agreement with the molecular dimensions obtained from various experimental methods.

The preferred conformations of the four oligomeric fragments of Rhamnogalacturonan II

Rhamnogalacturonan II (RG-II) is a structurally complex pectic mega-oligosaccharide that is released enzymatically from the primary cell wall of higher plants. RG-II contains 28 monosaccharide units (MW approximately equal to 6 KDa) which belong to 12 different families of glycosyl residues, including very unusual ones such as Kdo, Dha, aceric acid, and apiose. Eighteen different disaccharide segments can be identified, and so far the primary structure has not yet been determined. These monomeric units are arranged into four structurally well-defined oligosaccharide side chains, linked to a pectic backbone made up of 1,4-linked alpha-D-galactosyluronic acid residues. The specific attachment sites of these four side-chains on the pectic backbone remains to be elucidated. The present work presents a three-dimensional database of all the monosaccharide and disaccharide components of RG-II. The conformational behavior of D-Apif and L-AceAf monosaccharide has been assessed through computations performed with the molecular mechanics program MM3 using the flexible residue approach. For each furanosyl residue, energies of various envelope and twist conformers were systematically calculated as a function of the puckering parameters Q and phi. Energy minima are observed in both the Northern and Southern zones of the conformational wheel of each monosaccharide. As for the constituting segments, the conformational behaviour of 18 different disaccharides was evaluated using the flexible residue procedure of the MM3 molecular mechanics procedure. For each disaccharide, the adiabatic energy surface, along with the locations of the local energy minima and drawings of the conformations of each local minimum located in the energy maps have been established. The geometries of the minima and the potential energy surfaces of the different fragments were included in the database of the POLYS, a program for building oligo and polysaccharides. All these results were used for the generation, prior to a complete optimization, of the complete structure of each fragment of RG-II. It is shown that both A and B fragments are very flexible about the two sidechain glycosidic linkages which are closest to the backbone. The remaining part of the sidechain is rigid for the heavily branched A fragment, it is flexible for the more linear B fragment. The lowest energy conformer of each fragment results in good exposure of the hydroxyl groups of the apiosyl residues. Some possible implications of these features in boron complexation are presented.

Modelling of Congo red adsorption on the hydrophobic surface of cellulose using molecular dynamics

This study describes the interaction between crystalline cellulose and the direct dye Congo red (CR) using molecular dynamics simulations. A model of a microfibril corner of cellulose Iβ, exposing one hydrophobic (1 0 0) and the two hydrophilic (1 1 0) and (1 −1 0) surfaces was built. The energetic and geometric features of the dye adsorption were investigated at different temperatures following different initial positions of the CR. The relative positions and orientations of the CR with respect to the cellulose surface were unambiguously characterized using three translation (shift, slide, and rise) and three rotation (roll, tilt, and twist) parameters. Changes of twist and roll parameters showed that there was a tendency for the once adsorbed dye molecules, to become more planar and parallel to the cellulose surface. Several stable adsorption sites, translated laterally with respect to one another were obtained and adsorbed CR molecules were laid with their long axis either parallel or inclined by 50° with respect to the cellulose chain axis. Both vacuum and explicit water environments were considered. In this latter case, there was an increase in the energy of interaction between CR and cellulose and the adsorbed dye molecule behaved more rigidly than in vacuum case.