Supermolecular structure of cellulose/amylose blends prepared from aqueous NaOH solutions and effects of amylose on structural formation of cellulose from its solution

We previously proposed a mechanism for the structural formation of cellulose from its solution using a molecular dynamics (MD) simulation and suggested that the initial structure from its solution plays a critical role in determining its final structure. Structural changes in the van der Waals-associated cellulose molecular sheet as the initial structure were examined by MD simulation; the molecular sheet was found to be disordered due to maltohexaoses as an amylose model in terms of the hydrogen bonding system of cellulose. The structure and properties of cellulose/amylose blends prepared from an aqueous NaOH solution were examined experimentally by wide-angle X-ray diffraction and dynamic viscoelasticity measurements. The crystallinity of cellulose in the cellulose/amylose blend films was lower than that of cellulose film. The diffraction peaks of the cellulose/amylose blends were slightly shifted; specifically, () was shifted to a higher angle, and (1 1 0) and (0 2 0) were shifted to lower angles. These experimental results probably resulted from the disordered molecular sheet, as revealed by MD simulations.     

Thermal Stability of Apolipoprotein A-I in High-Density Lipoproteins by Molecular Dynamics

Apolipoprotein (apo) A-I is an unusually flexible protein whose lipid-associated structure is poorly understood. Thermal denaturation, which is used to measure the global helix stability of high-density lipoprotein (HDL)-associated apoA-I, provides no information about local helix stability. Here we report the use of temperature jump molecular dynamics (MD) simulations to scan the per-residue helix stability of apoA-I in phospholipid-rich HDL. When three 20 ns MD simulations were performed at 500 K on each of two particles created by MD simulations at 310 K, bilayers remained intact but expanded by 40%, and total apoA-I helicity decreased from 95% to 72%. Of significance, the conformations of the overlapping N- and C-terminal domains of apoA-I in the particles were unusually mobile, exposing hydrocarbon regions of the phospholipid to solvent; a lack of buried interhelical salt bridges in the terminal domains correlated with increased mobility. Nondenaturing gradient gels show that 40% expansion of the phospholipid surface of 100:2 particles by addition of palmitoyloleoylphosphatidylcholine exceeds the threshold of particle stability. As a unifying hypothesis, we propose that the terminal domains of apoA-I are phospholipid concentration-sensitive molecular triggers for fusion/remodeling of HDL particles. Since HDL remodeling is necessary for cholesterol transport, our model for remodeling has substantial biomedical implications.     

The Role of Cross-Chain Ionic Interactions for the Stability of Collagen Model Peptides

The contribution of ionic interactions to the stability of the collagen triple helix was studied using molecular dynamics (MD) simulations and biophysical methods. To this end, we examined the stability of a host-guest collagen model peptide, Ac-GPOGPOGPYGXOGPOGPO-NH₂, substituting KGE, KGD, EGK, and DGK for the YGX sequence. All-atom, implicit solvent MD simulations show that the fraction of cross-chain ionic interactions formed is different, with the most pronounced in the KGE and KGD sequences, and the least in the DGK sequence. To test whether the fraction of cross-chain ionic interactions correlates with the stability, experimental measurements of thermostability were done using differential scanning calorimetry and circular dichroism spectroscopy. It was found that the melting temperature is very similar for KGE and KGD peptides, whereas the EGK peptide has lower thermostability and the DGK peptide is the least thermostable. A novel, to our knowledge, computational protocol termed temperature-scan MD was applied to estimate the relative stabilities of the peptides from MD simulations. We found an excellent correlation between transition temperatures obtained from temperature-scan MD and those measured experimentally. These results suggest the importance of cross-chain ionic interactions for the stability of collagen triple helix and confirm the utility of MD simulations in predicting interactions and stability in this system.     

The Role of Cross-Chain Ionic Interactions for the Stability of Collagen Model Peptides

The contribution of ionic interactions to the stability of the collagen triple helix was studied using molecular dynamics (MD) simulations and biophysical methods. To this end, we examined the stability of a host-guest collagen model peptide, Ac-GPOGPOGPYGXOGPOGPO-NH₂, substituting KGE, KGD, EGK, and DGK for the YGX sequence. All-atom, implicit solvent MD simulations show that the fraction of cross-chain ionic interactions formed is different, with the most pronounced in the KGE and KGD sequences, and the least in the DGK sequence. To test whether the fraction of cross-chain ionic interactions correlates with the stability, experimental measurements of thermostability were done using differential scanning calorimetry and circular dichroism spectroscopy. It was found that the melting temperature is very similar for KGE and KGD peptides, whereas the EGK peptide has lower thermostability and the DGK peptide is the least thermostable. A novel, to our knowledge, computational protocol termed temperature-scan MD was applied to estimate the relative stabilities of the peptides from MD simulations. We found an excellent correlation between transition temperatures obtained from temperature-scan MD and those measured experimentally. These results suggest the importance of cross-chain ionic interactions for the stability of collagen triple helix and confirm the utility of MD simulations in predicting interactions and stability in this system.     

Chemoenzymatic syntheses of amylose-grafted chitin and chitosan

Amylose-grafted chitin and chitosan were synthesized by chemoenzymatic methods according to the following reaction manners. First, maltoheptaose was introduced to chitosan by a reductive amination using sodium cyanotrihydroborate in a mixed solvent of 1.0 mol/L aqueous acetic acid and methanol at room temperature to produce a maltoheptaose-grafted chitosan (1). The functionality of maltoheptaose to chitosan in 1 depended on reaction time. The phosphorylase-catalyzed enzymatic polymerization of R-D-glucose 1-phosphate was then performed from 1 to obtain amylose-grafted chitosan (2). Maltoheptaose-grafted chitin (3) was synthesized by N-acetylation of 1 using acetic anhydride in a mixed solvent of aqueous acetic acid and methanol. Then, synthesis of amylose-grafted chitin (4) was performed by the phosphorylase-catalyzed enzymatic polymerization under conditions the same as those for 2. The average DPs of amylose graft chains in 2 and 4 depended on the feed ratios of R-D-glucose 1-phosphate to maltoheptaose primers in 1 and 3.     

Nuclear overhauser effect spectroscopy (NOESY) detection of the specific interaction between substituents in cellulose and amylose triacetates

Nuclear Overhauser effect spectroscopy measurements on cellulose triacetate and on amylose triacetate with a mixing time of 500 ms, on the order of T₁, of acetyl protons, were performed to detect the specific through-space interaction between acetyl groups arising from their helix structures in solution. For cellulose triacetate, cross peaks were detected in CDCl₃ between acetyl proton signals at 3 and 6 positions on an anhydroglucose unit. In DMSO-d⁶, on the other hand, correlation peaks were observed not only between the 3 and 6 positions but also the 2 and 6 positions. For amylose triacetate, cross peaks were detected in CDCl₃ between the acetyl proton signals at the 2 and 6 positions. The through-space interaction of acetyl groups in cellulose triacetate and in amylose triacetate in solution was then interpreted based on their three-dimensional structures in solid state determined by x-ray crystallography. © 1994 John Wiley & Sons, Inc.     

Molecular dynamics simulation of the stability of a 22-residue α-helix in water and 30% trifluoroethanol

A molecular dynamics (MD) simulation was performed on the α-helix H8-HC5, the C-terminal part of myoglobin (residue 132–153), under periodic boundary conditions in two different solutions, water and water with 30% (v/v) 2,2,2-trifluoroethanol (TFE), at 300 K to investigate the stability of the helix. In both simulations, the initial configuration was a canonical right-handed α-helix. In the course of the MD trajectory in water (200 ps), the helix clearly destabilized and began to unfold after 100 ps. In the TFE solution, two stable parts of helical regions were observed after 70 ps of a 200-ps MD simulation, supporting the notion that TFE acts as a structure-forming solvent. © 1993 John Wiley & Sons, Inc.     

An alamethicin channel in a lipid bilayer: molecular dynamics simulations

We present the results of 2-ns molecular dynamics (MD) simulations of a hexameric bundle of Alm helices in a 1-palmitoyl-2-oleoylphosphatidylcholine bilayer. These simulations explore the dynamic properties of a model of a helix bundle channel in a complete phospholipid bilayer in an aqueous environment. We explore the stability and conformational dynamics of the bundle in a phospholipid bilayer. We also investigate the effect on bundle stability of the ionization state of the ring of Glu18 side chains. If all of the Glu18 side chains are ionised, the bundle is unstable; if none of the Glu18 side chains are ionized, the bundle is stable. pKA calculations suggest that either zero or one ionized Glu18 is present at neutral pH, correlating with the stable form of the helix bundle. The structural and dynamic properties of water in this model channel were examined. As in earlier in vacuo simulations (Breed et al., 1996 .Biophys. J. 70:1643-1661), the dipole moments of water molecules within the pore were aligned antiparallel to the helix dipoles. This contributes to the stability of the helix bundle.     

Conformational features of crystal-surface cellulose from higher plants

Native cellulose in higher plants forms crystalline fibrils a few nm across, with a substantial fraction of their glucan chains at the surface. The accepted crystal structures feature a flat-ribbon 21 helical chain conformation with every glucose residue locked to the next by hydrogen bonds from O-3' to O-5 and from O-2 to O-6'. Using solid-state NMR spectroscopy we show that the surface chains have a different C-6 conformation so that O-6 is not in the correct position for the hydrogen bond from O-2. We also present evidence consistent with a model in which alternate glucosyl residues are transiently or permanently twisted away from the flat-ribbon conformation of the chain, weakening the O-3' - 0-5 hydrogen bond. Previous molecular modelling and the modelling studies reported here indicate that this 'translational' chain conformation is energetically feasible and does not preclude binding of the surface chains to the interior chains, because the surface chains share the axial repeat distance of the 21 helix. Reduced intramolecular hydrogen bonding allows the surface chains to form more hydrogen bonds to external molecules in textiles, wood, paper and the living plant.     

Docking and molecular dynamics simulations of peroxisome proliferator activated receptors interacting with pan agonist sodelglitazar

PPAR (peroxisome proliferator-activated receptor) pan agonists play a critical role in treating metabolic diseases, especially the Type-2 diabetes mellitus (T2DM). GlaxoSmithKline's sodelglitazar (GW677954) is one of the potent PPAR pan agonists, which is currently being investigated in Phase II clinical trials for the treatment of T2DM and its complications. The present study was aimed at investigation into the effect of sodelglitazar at the binding pockets of PPARs. The Schrodinger Suite program (2009) was used for the molecular docking, while the GROMACS program used for the molecular dynamics (MD) simulations. The results thus obtained showed that sodelglitazar being docked well in the active site of PPARs. It was revealed by the MD simulations that the structures of the receptors remained quite stable during the simulations and that the important AF-2 helix showed less flexibility after binding with sodelglitazar. Also, it was observed that sodelglitazar could periodically form hydrogen bonds with the AF-2 helix of PPARs to stabilize the AF-2 helix in an active conformation. Our findings have confirmed that GlaxoSmithKline's sodelglitazar can activate the PPARs, which is quite consistent with the previous biological studies.     

On the mechanism of alpha-amylase.

Two inhibitors, acarbose and cyclodextrins (CD), were used to investigate the active site structure and function of barley alpha-amylase isozymes, AMY1 and AMY2. The hydrolysis of DP 4900-amylose, reduced (r) DP18-maltodextrin and maltoheptaose (catalysed by AMY1 and AMY2) was followed in the absence and in the presence of inhibitor. Without inhibitor, the highest activity was obtained with amylose, kcat/Km decreased 103-fold using rDP18-maltodextrin and 10(5) to 10(6)-fold using maltoheptaose as substrate. Acarbose is an uncompetitive inhibitor with inhibition constant (L1i) for amylose and maltodextrin in the micromolar range. Acarbose did not bind to the active site of the enzyme, but to a secondary site to give an abortive ESI complex. Only AMY2 has a second secondary binding site corresponding to an ESI2 complex. In contrast, acarbose is a mixed noncompetitive inhibitor of maltoheptaose hydrolysis. Consequently, in the presence of this oligosaccharide substrate, acarbose bound both to the active site and to a secondary binding site. alpha-CD inhibited the AMY1 and AMY2 catalysed hydrolysis of amylose, but was a very weak inhibitor compared to acarbose.beta- and gamma-CD are not inhibitors. These results are different from those obtained previously with PPA. However in AMY1, as already shown for amylases of animal and bacterial origin, in addition to the active site, one secondary carbohydrate binding site (s1) was necessary for activity whereas two secondary sites (s1 and s2) were required for the AMY2 activity. The first secondary site in both AMY1 and AMY2 was only functional when substrate was bound in the active site. This appears to be a general feature of the alpha-amylase family.     

Molecular dynamics investigations of the polysaccharide scleroglucan: first study on the triple helix structure

Explicit solvent molecular dynamics (MD) simulations on the triple helix of the polysaccharide Scleroglucan (Sclg) at two temperatures (273 and 300 K) were carried out. Owing to the complexity of the system, a united-atom force field, based on the properly modified GROMACS parameters, was adopted. To test these parameters for our system, MD simulations of the two disaccharidic units, representing the main chain and the side-chain linkages of the Sclg repeating unit, were performed and the results were compared with the literature data. The simulated triple helix of Sclg retained the main experimentally determined features of the polymer. The residence times of the solvent molecules at 273 and 300 K were analyzed. The results show that the more internal water molecules, interacting with the core of the Sclg triplex are not influenced substantially by changing the temperature, on the contrary the water molecules, interacting with the side-chain glucose residues show more significant differences. These data suggest that the more external water molecules, interacting with the side chain, play a major role in the conformational transition experimentally observed at low temperature.     

Molecular dynamics simulations of DNA curvature and flexibility: helix phasing and premelting

Recent studies of DNA axis curvature and flexibility based on molecular dynamics (MD) simulations on DNA are reviewed. The MD simulations are on DNA sequences up to 25 base pairs in length, including explicit consideration of counterions and waters in the computational model. MD studies are described for ApA steps, A-tracts, for sequences of A-tracts with helix phasing. In MD modeling, ApA steps and A-tracts in aqueous solution are essentially straight, relatively rigid, and exhibit the characteristic features associated with the B'-form of DNA. The results of MD modeling of A-tract oligonucleotides are validated by close accord with corresponding crystal structure results and nuclear magnetic resonance (NMR) nuclear Overhauser effect (NOE) and residual dipolar coupling (RDC) structures of d(CGCGAATTCGCG) and d(GGCAAAAAACGG). MD simulation successfully accounts for enhanced axis curvature in a set of three sequences with phased A-tracts studied to date. The primary origin of the axis curvature in the MD model is found at those pyrimidine/purine YpR "flexible hinge points" in a high roll, open hinge conformational substate. In the MD model of axis curvature in a DNA sequence with both phased A-tracts and YpR steps, the A-tracts appear to act as positioning elements that make the helix phasing more precise, and key YpR steps in the open hinge state serve as curvature elements. Our simulations on a phased A-tract sequence as a function of temperature show that the MD simulations exhibit a premelting transition in close accord with experiment, and predict that the mechanism involves a B'-to-B transition within A-tracts coupled with the prediction of a transition in key YpR steps from the high roll, open hinge, to a low roll, closed hinge substate. Diverse experimental observations on DNA curvature phenomena are examined in light of the MD model with no serious discrepancies. The collected MD results provide independent support for the "non-A-tract model" of DNA curvature. The "junction model" is indicated to be a special case of the non-A-tract model when there is a Y base at the 5' end of an A-tract. In accord with crystallography, the "ApA wedge model" is not supported by MD.     

A 1H-NMR and MD study of intramolecular hydrogen bonds in methyl beta-cellobioside.

The existence of an HO-3...O-5' intramolecular hydrogen bond in methyl beta-cellobioside in solution in Me2SO-d6 and H2O-CD3OD (4:1 w/w) was studied by 500-MHz 1H-NMR spectroscopy and MD simulations. Temperature coefficients for the chemical shift of the hydroxyl resonances in these solvents were determined and the rates of proton exchange in the latter solvent were obtained from NOE data. With H2O-CD3OD as the solvent, the HO-3...O-5' hydrogen bond was insignificant, but its presence in Me2SO-d6 was confirmed.     

A V7 conformation of dimethyl sulfoxide-amylose complex

X-ray investigation of a crystalline complex between dimethyl sulfoxide (DMSO) and amylose indicates a sevenfold helical structure having an orthogonal unit cell: a = 30.23 Å, b = 28.18 Å, and C = 7.91 Å (helix axis). This helical amylose results from precipitation of amylose in DMSO solutions under suitably dry conditions using toluene as a precipitating agent. Analyses show a molar ratio of 0.95:2:0.4:0.08 for DMSO, amylose glucose units, water, and toluene, respectively. DMSO and amylose combine in at least two crystalline conformations, sixfold and sevenfold helices. The amount of water present influences the resulting conformation. The existence of the sevenfold helical solid indicates that the amylose sevenfold helix has greater stability than previously recognized by conformational energy calculations. In general, potential energy calculations cannot be correct that ignore the influence of guest molecules and their effects upon the maltose residue conformation and that lead to a sixfold helix as the most probable structure.     

Helical conformation of amylose in aqueous solution. I. Viscosity measurements

Two amylose samples, amylose V (DP_w = 2300) and amylose HE 15 (a low-substituted hydroxyethylamylose, DP_W = 1600) were studied. The intrinsic viscosity of the polymers in aqueous solution was measured with regard to its dependence on the alkalinity (0 to 5MNaOH), the ionic strength (0 to 5M), and the temperature (0 to 75°C). Additionally, the temperature dependence of the viscosity of the amylose-iodine complex was measured. It was found that the two amylose samples show the same dependence on the studied parameters. Therefore, it was concluded that the conformation is unchanged by the hydroxyethylation in the present case. In a discussion, steric, hydrodynamic, and thermodynamic data on amylose in solution are compared with the corresponding data on cellulose and dextran. The comparison leads to the conclusion that amylose in a neutral solution must have a helical conformation, corresponding to the well-accepted rod conformation of cellulose. The helical conformation also explains several viritual anomalies in the behavior of amylose. The results of the experiments support the helix model for amylose. The conclusion of the whole work, therefore, is that the amylose molecule in neutral aqueous solution can be regarded as a random coil, built up by helical segments. The average number of monomers per segment exceeds 100. This value decreases with increasing alkalinity.     

Asymmetric stability among the transmembrane helices of lactose permease

Combining structure determinations from nuclear magnetic resonance (NMR) data and molecular dynamics simulations (MD) under the same environmental conditions revealed a startling asymmetry in the intrinsic conformational stability of secondary structure in the transmembrane domain of lactose permease (LacY). Eleven fragments, corresponding to transmembrane segments (TMs) of LacY, were synthesized, and their secondary structure in solution was determined by NMR. Eight of the TMs contained significant regions of helical structure. MD simulations, both in DMSO and in a DMPC bilayer, showed sites of local stability of helical structure in these TMs, punctuated by regions of conformational instability, in substantial agreement with the NMR data. Mapping the stable regions onto the crystal structure of LacY reveals a marked asymmetry, contrasting with the pseudosymmetry in the static structure: the secondary structure in the C-terminal half is more stable than in the N-terminal half. The relative stability of secondary structure is likely exploited in the transport mechanism of LacY. Residues supporting proton conduction are in more stable regions of secondary structure, while residues key to substrate binding are found in considerably unstable regions of secondary structure.     

Interpretation of H-NMR Experiments on the Orientation of the Transmembrane Helix WALP23 by Computer Simulations

Orientation, dynamics, and packing of transmembrane helical peptides are important determinants of membrane protein structure, dynamics, and function. Because it is difficult to investigate these aspects by studying real membrane proteins, model transmembrane helical peptides are widely used. NMR experiments provide information on both orientation and dynamics of peptides, but they require that motional models be interpreted. Different motional models yield different interpretations of quadrupolar splittings (QS) in terms of helix orientation and dynamics. Here, we use coarse-grained (CG) molecular dynamics (MD) simulations to investigate the behavior of a well-known model transmembrane peptide, WALP23, under different hydrophobic matching/mismatching conditions. We compare experimental ²H-NMR QS (directly measured in experiments), as well as helix tilt angle and azimuthal rotation (not directly measured), with CG MD simulation results. For QS, the agreement is significantly better than previously obtained with atomistic simulations, indicating that equilibrium sampling is more important than atomistic details for reproducing experimental QS. Calculations of helix orientation confirm that the interpretation of QS depends on the motional model used. Our simulations suggest that WALP23 can form dimers, which are more stable in an antiparallel arrangement. The origin of the preference for the antiparallel orientation lies not only in electrostatic interactions but also in better surface complementarity. In most cases, a mixture of monomers and antiparallel dimers provides better agreement with NMR data compared to the monomer and the parallel dimer. CG MD simulations allow predictions of helix orientation and dynamics and interpretation of QS data without requiring any assumption about the motional model.     

Two-dimensional correlation spectroscopy and principal component analysis studies of temperature-dependent IR spectra of cotton-cellulose

The FTIR spectra were measured for raw Uplands Sicala-V2 cotton fibers over a temperature range of 40-325 degrees C to explore the temperature-dependent changes in the hydrogen bonds of cellulose. These cotton-cellulose spectra exhibited complicated patterns in the 3800-2800 cm(-1) region and thus were analyzed by both the exploratory principal component analysis (PCA) and two-dimensional (2-D) correlation spectroscopy methods. The exploratory PCA showed that the spectra separate into two groups on the basis of thermal degradation of the cotton-cellulose and the consequent breakage of intersheet H-bonds present in its structure. Frequency variables, which strongly contributed to each principal component highlighted in its loadings plot, were linked to the frequencies assigned to vibrations of the OH groups involved in different kinds of H-bonds, as well as to vibrations of the CH groups. Deeper insights into reorganization of the temperature-dependent hydrogen bonding were obtained by 2-D correlation spectroscopy. Synchronous and asynchronous spectra were analyzed in the temperature ranges of 40 to 150 and 250 to 320 degrees C, the ranges indicated by PCA. Detailed band assignments of the OH stretching region and changes in the patterns of the hydrogen bonding network of the cotton-cellulose were proposed with the aid of the 2-D correlation spectroscopy analysis. Below 150 degrees C, distinctly different bands assigned to the less stable Ialpha and the more stable Ibeta interchain H-bonds O-6-H-6...O-3' were observed at about 3230 and 3270 cm(-1), respectively. Evaporation of water entrapped in the cellulose network was examined by means of the band at about 3610 cm(-1). The cooperativity of hydrogen bonds, which play a key role in the cellulose conformation, was monitored by frequencies assigned to intrachain H-bonds. It was possible to separate the frequencies assigned to the O-2-H-2...O-6 and O-3-H-3...O-5 intrachain H-bonds into two separate ranges, the spread of which was controlled by the cooperativity effect. The temperature dependence of the asynchronous spectra indicated that the less stable O-3-H-3...O-5 bonds gave rise to an absorption extending from 3300 to 3384 cm(-1), while the more stable O-2-H-2...O-6 bonds were characterized by the absorption between 3400 and 3470 cm(-1). The final breaking of the inter- and intrachain H-bonds, which occurs at the higher temperatures, was monitored by the asynchronous peaks at 3533 and 3590 cm(-1), respectively. On the basis of both the exploratory PCA and 2-D correlation spectroscopy investigations, it was possible to extract well-defined wavenumber ranges assigned to different kinds of intra- and interchain hydrogen bonds, as well as to the free OH groups of the cotton-cellulose.     

Interaction of the α2A domain of integrin with small collagen fragments

We here present a detailed study of the ligand-receptor interactions between single and triple-helical strands of collagen and the α2A domain of integrin (α2A), providing valuable new insights into the mechanisms and dynamics of collagen-integrin binding at a sub-molecular level. The occurrence of single and triple-helical strands of the collagen fragments was scrutinized with atom force microscopy (AFM) techniques. Strong interactions of the triple-stranded fragments comparable to those of collagen can only be detected for the 42mer triple-helical collagen-like peptide under study (which contains 42 amino acid residues per strand) by solid phase assays as well as by surface plasmon resonance (SPR) measurements. However, changes in NMR signals during titration and characteristic saturation transfer difference (STD) NMR signals are also detectable when α2A is added to a solution of the 21mer single-stranded collagen fragment. Molecular dynamics (MD) simulations employing different sets of force field parameters were applied to study the interaction between triple-helical or single-stranded collagen fragments with α2A. It is remarkable that even single-stranded collagen fragments can form various complexes with α2A showing significant differences in the complex stability with identical ligands. The results of MD simulations are in agreement with the signal alterations in our NMR experiments, which are indicative of the formation of weak complexes between single-stranded collagen and α2A in solution. These results provide useful information concerning possible interactions of α2A with small collagen fragments that are of relevance to the design of novel therapeutic A-domain inhibitors.