4C1 Chair Research Articles

Reaction Mechanism of Glycoside Hydrolase Family 116 Utilizes Perpendicular Protonation.

Retaining glycoside hydrolases use acid/base catalysis with an enzymatic acid/base protonating the glycosidic bond oxygen to facilitate leaving-group departure alongside attack by a catalytic nucleophile to form a covalent intermediate. Generally, this acid/base protonates the oxygen laterally with respect to the sugar ring, which places the catalytic acid/base and nucleophile carboxylates within about 4.5-6.5 Å of each other. However, in glycoside hydrolase (GH) family 116, including disease-related human acid β-glucosidase 2 (GBA2), the distance between the catalytic acid/base and the nucleophile is around 8 Å (PDB: 5BVU) and the catalytic acid/base appears to be above the plane of the pyranose ring, rather than being lateral to that plane, which could have catalytic consequences. However, no structure of an enzyme-substrate complex is available for this GH family. Here, we report the structures of Thermoanaerobacterium xylanolyticum β-glucosidase (TxGH116) D593N acid/base mutant in complexes with cellobiose and laminaribiose and its catalytic mechanism. We confirm that the amide hydrogen bonding to the glycosidic oxygen is in a perpendicular rather than lateral orientation. Quantum mechanics/molecular mechanics (QM/MM) simulations of the glycosylation half-reaction in wild-type TxGH116 indicate that the substrate binds with the nonreducing glucose residue in an unusual relaxed 4C1 chair at the -1 subsite. Nevertheless, the reaction can still proceed through a 4H3 half-chair transition state, as in classical retaining β-glucosidases, as the catalytic acid D593 protonates the perpendicular electron pair. The glucose C6OH is locked in a gauche, trans orientation with respect to the C5-O5 and C4-C5 bonds to facilitate perpendicular protonation. These data imply a unique protonation trajectory in Clan-O glycoside hydrolases, which has strong implications for the design of inhibitors specific to either lateral protonators, such as human GBA1, or perpendicular protonators, such as human GBA2.

Enzymatic C4-Epimerization of UDP-Glucuronic Acid: Precisely Steered Rotation of a Transient 4-Keto Intermediate for an Inverted Reaction without Decarboxylation.

UDP-glucuronic acid (UDP-GlcA) 4-epimerase illustrates an important problem regarding enzyme catalysis: balancing conformational flexibility with precise positioning. The enzyme coordinates the C4-oxidation of the substrate by NAD+ and rotation of a decarboxylation-prone β-keto acid intermediate in the active site, enabling stereoinverting reduction of the keto group by NADH. We reveal the elusive rotational landscape of the 4-keto intermediate. Distortion of the sugar ring into boat conformations induces torsional mobility in the enzyme's binding pocket. The rotational endpoints show that the 4-keto sugar has an undistorted 4 C1 chair conformation. The equatorially placed carboxylate group disfavors decarboxylation of the 4-keto sugar. Epimerase variants lead to decarboxylation upon removal of the binding interactions with the carboxylate group in the opposite rotational isomer of the substrate. Substitutions R185A/D convert the epimerase into UDP-xylose synthases that decarboxylate UDP-GlcA in stereospecific, configuration-retaining reactions.

Impact of Polarization on the Ring Puckering Dynamics of Hexose Monosaccharides.

Analysis of crystal structures of hexose monosaccharides α-d-mannose (α-MAN), β-d-mannose (β-MAN), α-d-glucose (α-GLC), β-d-glucose (β-GLC), α-d-galactose (α-GAL), β-d-galactose (β-GAL), α-d-altrose (α-ALT), β-d-altrose (β-ALT), α-d-idose (α-IDO), and β-d-idose (β-IDO) reveals that the monosaccharide ring adopts multiple ring conformations. These ring conformations can be broadly classified as chair, half-chair, envelope, boat, and skew-boat conformations. The ability of the monosaccharide ring to adopt multiple conformations has been closely tied with their bioactivity. However, it has been difficult to capture the dynamic information of these conformations from experimental studies. Even from simulations, capturing these different conformations is challenging because of the energy barriers involved in the transitions between the stable 4C1 and 1C4 chair forms. In this study, we analyze the influence of the polarizable force field on the ring dynamics of five major types of unsubstituted aldohexoses─glucose, mannose, galactose, altrose, and idose─and their anomers. We simulate microsecond trajectories to capture the influence of the CHARMM36 additive and polarizable carbohydrate force fields on the ring dynamics. The microsecond trajectories allow us to comment on the issues associated with equilibrium molecular dynamics simulations. Further, we use the extended system adaptive biasing force (eABF) method to compare the conformational sampling efficiencies of the additive and polarizable force fields. Our studies reveal that inclusion of polarization enhances the sampling of ring conformations and lowers the energy barriers between the 4C1 and 1C4 conformations. Overall, the CHARMM36 additive force field is observed to be rigid and favor the 4C1 conformations. Although the inclusion of polarizability results in enhancing ring flexibility, we observe sampling that does not agree with experimental results, warranting a revision of the polarizable Drude parameters.

The carbohydrate glycosylphosphatidylinositol anchor chain under mechanical stress

Carbohydrates have quite complicated micro heterogenic structure which may undergo different structural transitions. Due to their extreme flexibility it is very difficult to investigate such structural changes experimentally. In these studies we want to predict what structural and conformational changes are possible in the carbohydrate glycosylphosphatidylinositol anchor chain (GPI): the tetrasaccharide with the unique sequence Man-α(1 → 2)-Man-α(1 → 6)-Man-α(1 → 4)-GlcN-α. This is a very important biomolecule associated with the processes of transmitting various types of signals in the cells of living organisms. In order to investigate conformational and structural changes in GPI we use in these studies the theoretical Enforced Geometry Optimization (EGO) method. In this method a molecule is exposed to a mechanical stress caused by external forces applied to selected atoms. It turned out that under external stretching forces the mannopyranose unit can change its 4C1 chair conformation into three different forms: 1S3, oS2 and B2,5. The initial 4C1 glucosamine ring can transit into the twisted boat 1S3 and the boat Bo,3 conformations. The obtained results confirm the high flexibility of the GPI anchor sugar chain.

Role of Enzyme and Active Site Conformational Dynamics in the Catalysis by α-Amylase Explored with QM/MM Molecular Dynamics.

We assessed enzyme:substrate conformational dynamics and the rate-limiting glycosylation step of a human pancreatic α-amylase:maltopentose complex. Microsecond molecular dynamics simulations suggested that the distance of the catalytic Asp197 nucleophile to the anomeric carbon of the buried glucoside is responsible for most of the enzyme active site fluctuations and that both Asp197 and Asp300 interact the most with the buried glucoside unit. The buried glucoside binds either in a 4C1 chair or 2SO skew conformations, both of which can change to TS-like conformations characteristic of retaining glucosidases. Starting from four distinct enzyme:substrate complexes, umbrella sampling quantum mechanics/molecular mechanics simulations (converged within less than 1 kcal·mol-1 within a total simulation time of 1.6 ns) indicated that the reaction occurrs with a Gibbs barrier of 13.9 kcal·mol -1, in one asynchronous concerted step encompassing an acid-base reaction with Glu233 followed by a loose SN2-like nucleophilic substitution by the Asp197. The transition state is characterized by a 2H3 half-chair conformation of the buried glucoside that quickly changes to the E3 envelope conformation preceding the attack of the anomeric carbon by the Asp197 nucleophile. Thermodynamic analysis of the reaction supported that a water molecule tightly hydrogen bonded to the glycosidic oxygen of the substrate at the reactant state (∼1.6 Å) forms a short hydrogen bond with Glu233 at the transition state (∼1.7 Å) and lowers the Gibbs barrier in over 5 kcal·mol-1. The resulting Asp197-glycosyl was mostly found in the 4C1 conformation, although the more endergonic B3,O conformation was also observed. Altogether, the combination of short distances for the acid-base reaction with the Glu233 and for the nucleophilic attack by the Asp197 nucleophile and the availability of water within hydrogen bonding distance of the glycosidic oxygen provides a reliable criteria to identify reactive conformations of α-amylase complexes.

D-Mannosamine hydrochloride (2-amino-2-deoxy-D-mannose hydrochloride): ionic hydrogen bonding in saccharides involving chloride and aminium ions.

D-Mannosamine hydrochloride (2-amino-2-deoxy-D-mannose hydrochloride), C6H14NO5+·Cl-, (I), crystallized from a methanol/ethyl acetate/n-hexane solvent mixture at room temperature in a 4C1 chair conformation that is slightly distorted towards the C3,O5B form. A comparison of the structural parameters of (I) with the corresponding parameters in α-D-glucosamine hydrochloride, (II), and β-D-galactosamine hydrochloride, (III)/(III'), was undertaken to evaluate the effects of ionic hydrogen bonding on structural properties. Three types of ionic hydrogen bonds are present in the crystals of (I)-(III)/(III'), i.e. N+-H...O, N+-H...Cl-, and O-H...Cl-. The exocyclic structural parameters in (I), (II), and (III)/(III') appear to be most influenced by this bonding, especially the exocyclic hydroxy groups, which adopt eclipsed conformations enabled by ionic hydrogen bonding to the chloride anion. Anomeric disorder was observed in crystals of (I), with an α:β ratio of 37:63. However, anomeric configuration appears to exert minimal structural effects; that is, bond lengths, bond angles, and torsion angles are essentially identical in both anomers. The observed disorder at the anomeric C atom of (I) appears to be caused by the presence of the chloride anion and atom O3 or O4 in proximal voids, which provide opportunities for hydrogen bonding to atom O1 in both axial and equatorial orientations.

Conformational preferences of triantennary and tetraantennary hybrid N-glycans in aqueous solution: Insights from 20 μs long atomistic molecular dynamic simulations

In the current study, we have investigated the conformational dynamics of a triantennary (N-glycan1) and tetraantennary (N-glycan2) hybrid N-glycans found on the surface of the HIV glycoprotein using 20 μs long all-atom molecular dynamics (MD) simulations. The main objective of the present study is to elucidate the influence of adding a complex branch on the overall glycan structural dynamics. Our investigation suggests that the average RMSD value increases when a complex branch is added to N-glycan1. However, the RMSD distribution is relatively wider in the case of N-glycan1 compared to N-glycan2, which indicates that multiple complex branches restrict the conformational variability of glycans. A similar observation is obtained from the principal component analysis of both glycans. All the puckering states (4C1 to 1C4) of each monosaccharide except mannose are sampled in our simulations, although the 4C1 chair form is energetically more favorable than 1C4. In N-glycan1, the 1–6 linkage in the mannose branch [Man(9)-α(1-6)-Man(5)] stays in the gauche-gauche cluster, whereas it moves towards trans-gauche in N-glycan2. For both glycans, mannose branches are more flexible than the complex branches, and adding a complex branch does not influence the dynamics of the mannose branches. We have noticed that the end-to-end distance of the complex branch shortens by ∼ 10 Å in the presence of another complex branch. This suggests that in the presence of an additional complex branch, the other complex branch adopts a close folded structure. All these conformational changes involve the selective formation of inter-residue and water-mediated hydrogen-bond networks.

Uridine-Derived 4-Aminophenyl 1-Thioglucosides: DFT Optimized FMO, ADME, and Antiviral Activities Study

The biological roles of carbohydrates, especially their synthetic glycoconjugates, are at the forefront of research, and their applications and research are in the development of an impressive area of carbohydrate-derived vaccines. Considering this, several thioglucosides 2-6 conjugated with para-aminophenyl and uridine moieties are investigated for their possible application as hepatitis antiviral applications. Geometry optimization of these uridine thioglucosides by density functional theory (DFT) indicated that their glucopyranose rings are in 4C1 chair conformation with β-glucosidic linkage. Frontier molecular orbital (FMO), molecular electrostatic potential (MEP), lower hardness, and softness calculations indicated their higher electrophilic nature. Solubility and ADMET studies indicated that the uridine thioglucosides might be safer drugs to use. With these encouraging results, molecular docking of 2-6 was conducted with hepatitis C virus (HCV, 3su4) and hepatitis B virus (HBV, 5e0i) proteases where the binding affinities for 2-6 were found higher than the standard anti-hepatitis drug sofosbuvir. Overall, the thioglucosides showed a better binding affinity with HBV protease (5e0i) than the HCV protease (3su4), indicating their prospect against these viruses.

Pyranose Ring Puckering Thermodynamics for Glycan Monosaccharides Associated with Vertebrate Proteins.

The conformational properties of carbohydrates can contribute to protein structure directly through covalent conjugation in the cases of glycoproteins and proteoglycans and indirectly in the case of transmembrane proteins embedded in glycolipid-containing bilayers. However, there continue to be significant challenges associated with experimental structural biology of such carbohydrate-containing systems. All-atom explicit-solvent molecular dynamics simulations provide a direct atomic resolution view of biomolecular dynamics and thermodynamics, but the accuracy of the results depends on the quality of the force field parametrization used in the simulations. A key determinant of the conformational properties of carbohydrates is ring puckering. Here, we applied extended system adaptive biasing force (eABF) all-atom explicit-solvent molecular dynamics simulations to characterize the ring puckering thermodynamics of the ten common pyranose monosaccharides found in vertebrate biology (as represented by the CHARMM carbohydrate force field). The results, along with those for idose, demonstrate that the CHARMM force field reliably models ring puckering across this diverse set of molecules, including accurately capturing the subtle balance between 4C1 and 1C4 chair conformations in the cases of iduronate and of idose. This suggests the broad applicability of the force field for accurate modeling of carbohydrate-containing vertebrate biomolecules such as glycoproteins, proteoglycans, and glycolipids.

Multidimensional Free Energy and Accelerated Quantum Library Methods Provide a Gateway to Glycoenzyme Conformational, Electronic, and Reaction Mechanisms.

Enzyme reactions are complex to simulate accurately, and none more so than glycoenzymes (glycosyltransferase and glycosidases). A rigorous sampling of the protein frame and the conformationally plural carbohydrate substrate coupled with an unbiased treatment of the electron dynamics is needed to discover the true reaction landscapes. Here, we demonstrate the effectiveness of two computational methods ported in libraries that we have developed. The first is a flat histogram free energy method called FEARCF capable of multidimensional sampling and rapidly converging to a complete coverage of phase space. The second, the Quantum Supercharger Library (QSL), is a method that accelerates the computation of the ab initio electronic wave function as well as the integral derivatives on graphical processing units (GPUs). These QSL accelerated computations form the core components needed for direct quantum dynamics and QM/MM dynamics when coupled with legacy codes such as GAMESS and NWCHEM, making state of the art hyper-parallel electronic computations in chemistry and chemical biology possible. The combination of QSL (acceleration of ab initio QM computation) and FEARCF (multidimensional hyper-parallel reaction dynamics) makes the simulation of ab initio QM/MM reaction dynamics of enzyme catalysis feasible. Enzymes that process carbohydrates pose an added challenge as their pyranose ring substrates span multidimensional conformational space whose sampling is an intimate function of the catalytic mechanism. Here, we use the pairing of FEARCF and QSL to simulate the catalytic effect of the glycoenzyme β-N-acetylglucosamine transferase (OGT). The reaction mechanism is discovered from a variable three bond reaction surface using SCCDFTB. The role of the OGT in distorting the pyranose ring of β-N-acetylglucosamine (GlcNAc) away from the equilibrium 4C1 chair conformation toward the E3 envelope needed for the transition state is discovered from its pucker free energy hypersurfaces (or free energy volume, FEV). A complete GlcNAc ring pucker HF 6-31g FEV is constructed from ab initio QM dynamics in vacuum and ab initio QM/MM dynamics in the OGT catalytic domain. The OGT is shown to clearly lower the pathway toward the transition state E3 ring conformer as well as stabilize it by 1.63 kcal/mol. Illustrated here is the use of QSL accelerated ab initio QM/MM dynamics that thoroughly explores carbohydrate catalyzed reactions through a FEARCF multidimensional sampling of the interdependence between reaction and conformational space. This demonstrates how experimentally inaccessible molecular and electronic mechanisms that underpin enzyme catalysis can be discovered by directly modeling the dynamics of these complex reactions.

Methyl β-lactoside [methyl β-D-galactopyranosyl-(1→4)-β-D-glucopyranoside] monohydrate: a solvomorphism study.

Methyl β-lactoside [methyl β-D-galactopyranosyl-(1→4)-β-D-glucopyranoside] monohydrate, C13H24O11·H2O, (I), was obtained via spontaneous transformation of methyl β-lactoside methanol solvate, (II), during air-drying. Cremer-Pople puckering parameters indicate that the β-D-Galp (β-D-galactopyranosyl) and β-D-Glcp (β-D-glucopyranosyl) rings in (I) adopt slightly distorted 4C1 chair conformations, with the former distorted towards a boat form (BC1,C4) and the latter towards a twist-boat form (O5SC2). Puckering parameters for (I) and (II) indicate that the conformation of the βGalp ring is slightly more affected than the βGlcp ring by the solvomorphism. Conformations of the terminal O-glycosidic linkages in (I) and (II) are virtually identical, whereas those of the internal O-glycosidic linkage show torsion-angle changes of 6° in both C-O bonds. The exocyclic hydroxymethyl group in the βGalp residue adopts a gt conformation (C4' anti to O6') in both (I) and (II), whereas that in the βGlcp residue adopts a gg (gauche-gauche) conformation (H5 anti to O6) in (II) and a gt (gauche-trans) conformation (C4 anti to O6) in (I). The latter conformational change is critical to the solvomorphism in that it allows water to participate in three hydrogen bonds in (I) as opposed to only two hydrogen bonds in (II), potentially producing a more energetically stable structure for (I) than for (II). Visual inspection of the crystalline lattice of (II) reveals channels in which methanol solvent resides and through which solvent might exchange during solvomorphism. These channels are less apparent in the crystalline lattice of (I).

Isopropyl 3-deoxy-α-D-ribo-hexopyranoside (isopropyl 3-deoxy-α-D-glucopyranoside): evaluating trends in structural parameters.

Isopropyl 3-deoxy-α-D-ribo-hexopyranoside (isopropyl 3-deoxy-α-D-glucopyranoside), C9H18O5, (I), crystallizes from a methanol-ethyl acetate solvent mixture at room temperature in a 4C1 chair conformation that is slightly distorted towards the C5SC1 twist-boat form. A comparison of the structural parameters in (I), methyl α-D-glucopyranoside, (II), α-D-glucopyranosyl-(1→4)-D-glucitol (maltitol), (III), and 3-deoxy-α-D-ribo-hexopyranose (3-deoxy-α-D-glucopyranose), (IV), shows that most endocyclic and exocyclic bond lengths, valence bond angles and torsion angles in the aldohexopyranosyl rings are more affected by anomeric configuration, aglycone structure and/or the conformation of exocyclic substituents, such as hydroxymethyl groups, than by monodeoxygenation at C3. The structural effects observed in the crystal structures of (I)-(IV) were confirmed though density functional theory (DFT) calculations in computed structures (I)c-(IV)c. Exocyclic hydroxymethyl groups adopt the gauche-gauche (gg) conformation (H5 anti to O6) in (I) and (III), and the gauche-trans (gt) conformation (C4 anti to O6) in (II) and (IV). The O-glycoside linkage conformations in (I) and (III) resemble those observed in disaccharides containing β-(1→4) linkages.

Conformational Analysis of Heparin-Analogue Pentasaccharides by Nuclear Magnetic Resonance Spectroscopy and Molecular Dynamics Simulations.

Elucidation and improvement of the blood coagulant properties of heparin are the focus of intense research. In this study, we performed conformational analysis using nuclear magnetic resonance (NMR) spectroscopy and molecular dynamics (MD) simulations on the heparin pentasaccharide analogue idraparinux, its disulfonatomethyl analogue, which features a slightly improved blood coagulation property, and a trisulfonatomethyl analogue, in which the activity has been totally abolished. As the ring conformation of the G subunit has been suggested as a major determinant of the biological properties, we analyzed the sugar ring conformations and dynamics of the interglycosidic linkages. We found that the conformation of the G ring is dominated by the 2SO skewed boat next to the 1C4 chair in all three derivatives. Both the thermodynamics and the kinetics of the conformational states were found to be highly similar in the three derivatives. Molecular kinetic analysis showed that the 2SO skewed boat state of the G ring is equally favorable in the three analogues, resulting in similar 2SO populations. Also, the transition kinetics from the 1C4 chair to the 2SO skewed boat was found to be comparable in the derivatives, which indicates a similar energy barrier between the two states of the G subunit. We also identified a slower conformational transition between the dominant 4C1 chair and the boat conformations on the E subunit. Both G and E ring flips are also accompanied by changes along the interglycosidic linkages, which take place highly synchronously with the ring flips. These findings indicate that conformational plasticity of the G ring and the dominance of the 2SO skewed boat populations do not necessarily warrant the biological activity of the derivatives and hence the impact of other factors also needs to be considered.

QM/MM Simulations of Enzymatic Hydrolysis of Cellulose: Probing the Viability of an Endocyclic Mechanism for an Inverting Cellulase.

Glycoside hydrolases (GH) cleave carbohydrate glycosidic bonds and play pivotal roles in living organisms and in many industrial processes. Unlike acid-catalyzed hydrolysis of carbohydrates in solution, which can occur either via cyclic or acyclic oxocarbenium-like transition states, it is widely accepted that GH-catalyzed hydrolysis proceeds via a general acid mechanism involving a cyclic oxocarbenium-like transition state with protonation of the glycosidic oxygen. The GH45 subfamily C inverting endoglucanase from Phanerochaete chrysosporium (PcCel45A) defies the classical inverting mechanism as its crystal structure conspicuously lacks a general Asp or Glu base residue. Instead, PcCel45A has an Asn residue, a notoriously weak base in solution, as one of its catalytic residues at position 92. Moreover, unlike other inverting GHs, the relative position of the catalytic residues in PcCel45A impairs the proton abstraction from the nucleophilic water that attacks the anomeric carbon, a key step in the classical mechanism. Here, we investigate the viability of an endocyclic mechanism for PcCel45A using hybrid quantum mechanics/molecular mechanics (QM/MM) simulations, with the QM region treated with the self-consistent-charge density-functional tight-binding level of theory. In this mechanism, an acyclic oxocarbenium-like transition state is stabilized leading to the opening of the glucopyranose ring and formation of an unstable acyclic hemiacetal that can be readily decomposed into hydrolysis product. In silico characterization of the Michaelis complex shows that PcCel45A significantly restrains the sugar ring to the 4C1 chair conformation at the −1 subsite of the substrate binding cleft, in contrast to the classical exocyclic mechanism in which ring puckering is critical. We also show that PcCel45A provides an environment where the catalytic Asn92 residue in its standard amide form participates in a cooperative hydrogen bond network resulting in its increased nucleophilicity due to an increased negative charge on the oxygen atom. Our results for PcCel45A suggest that carbohydrate hydrolysis catalyzed by GHs may take an alternative route from the classical mechanism.

Oxidative Addition of α‐Glycosyl Halides to a Platinum(0) Olefin Complex: Stereochemistry of Pt−C Bond Formation

AbstractThe oxidative addition of α‐glucosyl and α‐galactosyl halides to the platinum(0) complex [Pt(2,9‐dimethyl‐1,10‐phenanthroline)(ethene)] has been investigated. In all cases the main reaction product is the five‐coordinate Pt(II) complex containing in axial position an α‐glycosyl fragment in the unusual 1C4 conformation. In the case of glucose, a minor product could be identified as the β isomer, which retains the typical 4C1 chair. The single crystal structures of two diastereomers are compared. The outcome of the reaction does not depend on the nature of the halide and is only slightly affected by the solvent. The addition mechanism is discussed, also in the light of literature data. Experimental data, along with DFT calculations, point towards a radical chain mechanism.

The conformation of the idopyranose ring revisited: How subtle O-substituent induced changes can be deduced from vicinal 1H-NMR coupling constants

The idopyranose ring plays a pivotal role in the conformational, dynamical, and intermolecular binding aspects of glycosaminoglycans like heparin and dermatan sulfate and it was early on assigned a role in the Sugar Code governing biological recognition processes. There is consensus that next to the two canonical 1C4 and 4C1 chair conformations, the conformational space accessible to the idopyranose ring entails a 2SO skew-boat conformation, but the equilibrium between these three ring puckers has evaded satisfactory quantification. In this study a meta-analysis of X-ray solid-state data and vicinal NMR coupling constants is presented, based on the Truncated Fourier Puckering (TFP) formalism and the generalized Karplus (CAGPLUS) equation. This approach yields a model-free, granular and consistent reckoning of 159 idopyranose solution puckering equilibria studied by NMR and allows us to reproduce the involved 636 NMR vicinal couplings with an overall residual RMS(Jobs-Jcalc) of 0.184 Hz. Our analyses show that for all ring systems examined, the idopyranosyl chair conformations take up the same ring pucker irrespective of the ring substituent pattern or a vast variety in experimental conditions. Instead, it is the (skew-)boat conformation that adapts to the substitution pattern of the idopyranose ring or a specific sulfation pattern of neighboring saccharides. All idopyranose rings are involved in conformational equilibria that subsume the aforementioned conformers which turn out to differ only a few kJ/mole in conformational energy. Thus, the plasticity and flexibility of idopyranose remains intact under practically all circumstances and, as the glycosidic linkages in heparin are considered to be relatively stiff, the iduronic moiety functions as the linchpin of heparin flexibility thereby being rather a “space(r)” than a “letter” in the alleged Sugar Code alphabet.

The effects of HHP (high hydrostatic pressure) on the interchain interaction and the conformation of amylopectin and double-amylose molecules

Starch is an important resource in nature, and HHP (high hydrostatic pressure) is one of the most important physical modification technologies. In this study, molecular dynamics simulation was used to explore the interchain interaction and the changes of molecule conformations of amylopectin and double-amylose helix at atomic level in different pressure. The results shown that, firstly, high pressure increased the content of 4C1 chair conformation, decreased the RMSD (root mean square deviations) and RMSF (root mean square fluctuation), made molecules more stable. Secondly, high pressure increased the interchain VDW (Van der Waals) and electrostatic forces, then caused the decreases of the interchain distances and surface area of both amylopectin and double-amylose, made molecules more compact. Thirdly, high pressure decreased the intramolecular hydrogen bonds, increased the molecule-solvent hydrogen bonds. These findings can explain some existing experimental phenomena from the atomic level, meanwhile, it may also provide importance reference value for using of HHP in starch processing and the studies of starch granule structure.

Synthesis and Photochromic Properties of Azobenzene‐Derived Glycomacrolactones

Reversible photocontrol of glycosides and glycoconjugates structures is a very attractive approach to modulate, in a spatiotemporal way, the various properties and biological activities of carbohydrates. We have synthesized three new azobenzene‐derived glycomacrolactones from thioglycopyranosides. The synthesized cyclic glycoazobenzenes can be reversibly photoisomerized between E and Z isomers with high fatigue resistance. A 1H NMR spectroscopic study shows that E → Z isomerization of glycomacrocycles induces large conformational change of the macrocyclic structures, without changing sugar 4C1 chair conformation. The Z‐glycoazobenzenes can be thermally converted back to the E‐isomers. Interestingly, these 16 to 17‐membered Z‐glycomacrolactones display higher thermal stability than the reported macrocyclic azobenzenes, the half‐life varying from 37 to 72 days. The excellent photoswitching property and bistability of the synthesized glycoazobenzenes open a new opportunity for the convergent synthesis of diastereomers of glycomacrocycles. Furthermore, chiroptical properties have been observed for both E and Z glycomacrolactones. The geometry of different isomers of macrocycles has been optimized with DFT calculations. Theoretical CD spectra obtained by TD‐DFT suggest that the E and Z glycomacrocycles adopt preferentially (P) helical structure for the azobenzene moiety.

Development of Mannopyranoside Therapeutics against Adherent-Invasive Escherichia coli Infections.

Preventing bacterial adhesion to host cells is a provocative and alternative approach to traditional antibiotic treatments given the increasing microbial resistance. A brief overview of common antibiotic treatments is described in light of their respective resistance and remaining susceptibility. This strategy has been seriously considered in the context of adherent-invasive infections in Crohn's disease and urinary tract infections in particular. The adhesions of various pathogenic Escherichia coli strains to host cells are primarily mediated through carbohydrate-protein interactions involving bacterial organelles called fimbriae that can recognize specific glycoconjugate receptors on host cells. Of particular interest are the FimH and PapG fimbriae, which bind to mannosylated glycoproteins and glycolipids of the galabiose series, respectively. Therefore, blocking FimH- and PapG-mediated bacterial adhesion to uroepithelial cells by high-affinity carbohydrate antagonists constitutes a challenging therapeutic target of high interest. This is of particular interest since bacterial adhesion to host cells is a parameter unlikely to be the subject of bacterial mutations without affecting the carbohydrate ligand binding interactions at the basis of the recognition and infection processes. To date, there have been several families of potent FimH antagonists that include natural O-linked as well as unnatural analogues of α-d-mannopyranosides. These observations led to a thorough understanding of the intimate binding site interactions that helped to reveal the so-called "tyrosine gate mechanism" at the origin of the strong necessary interactions with sugar-possessing hydrophobic aglycones. By modification of the aglycones of single monosaccharidic d-mannopyranosides, it was possible to replace the natural complex oligomannoside structure by simpler ones. An appealing and successful series of analogues have been disclosed, including nanomolecular architectures such as dendrimers, polymers, and liposomes. In addition, the data were compared to the above multivalent architectures and confirmed the possibility of working with small sugar candidates. This Account primarily concentrates on the most promising types of FimH inhibitors belonging to the family of α-C-linked mannopyranosides. However, one of the drawbacks associated with C-mannopyranosides has been that they were believed to be in the inverted chair conformation, which is obviously not recognized by the E. coli FimH. To decipher this situation, various synthetic approaches, conformational aspects, and restrictions are discussed using molecular modeling, high-field NMR spectroscopy, and X-ray analysis. These combined techniques pointed to the fact that several α-C-linked mannopyranosides do exist in the required 4C1 chair conformation. Ultimately, recent findings in this growing field of interest culminated in the identification of drug candidates that have reached clinical phase I.

Hydrolysis and Transglycosylation Transition States of Glycoside Hydrolase Family 3 β-Glucosidases Differ in Charge and Puckering Conformation

β-Glucosidases (βgls) from glycoside hydrolase family 3 play an important role in biomass degradation by catalyzing cellobiose hydrolysis. However, the hydrolysis rate decreases when the glucose product or another cellobiose competes with water to form oligosaccharides in a reaction called transglycosylation. Both reactions involve proton transfer to the acid/base residue and nucleophilic attack on the glycosyl-enzyme intermediate. To gain a deeper understanding of these competing reactions, quantum mechanics/molecular mechanics calculations were performed. Although both reactions are exothermic and have similar free-energy barriers (∼18 kcal/mol), the transition-state (TS) characteristics are different. The glycosyl-water bond is nearly formed in the hydrolysis TS, leading to reduced ionic character and a 4C1 chair conformation. The transglycosylation TS is more positively charged and adopts the 4H3 half-chair conformation because bond formation is less advanced. Water interacts solely with acid/base residue E441, though the long distance between them (2.1 Å) suggests that E441 does not activate water for nucleophilic attack. In comparison, a glucose acceptor has a lower deprotonation enthalpy and hydrogen bonds to E441 (1.6 Å) as well as to Y204, R169, and R67. Knowledge of these factors that are relevant to TS formation and stability is valuable for engineering βgls with enhanced hydrolytic activity for industrial applications.