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Dorst, K. & Widmalm, G. (2024). Conformational Preferences at the Glycosidic Linkage of Saccharides in Solution as Deduced from NMR Experiments and MD Simulations: Comparison to Crystal Structures. Chemistry - A European Journal
Open this publication in new window or tab >>Conformational Preferences at the Glycosidic Linkage of Saccharides in Solution as Deduced from NMR Experiments and MD Simulations: Comparison to Crystal Structures
2024 (English)In: Chemistry - A European Journal, ISSN 0947-6539, E-ISSN 1521-3765Article in journal (Refereed) Epub ahead of print
Abstract [en]

Glycans are central to information content and regulation in biological systems. These carbohydrate molecules are active either as oligo- or polysaccharides, often in the form of glycoconjugates. The monosaccharide entities are joined by glycosidic linkages and stereochemical arrangements are of utmost importance in determining conformation and flexibility of saccharides. The conformational preferences and population distributions at the glycosidic torsion angles phi and psi have been investigated for O-methyl glycosides of three disaccharides where the substitution takes place at a secondary alcohol, viz., in alpha-l-Fucp-(1 -> 3)-beta-d-Glcp-OMe, alpha-l-Fucp-(1 -> 3)-alpha-d-Galp-OMe and alpha-d-Glcp-(1 -> 4)-alpha-d-Galp-OMe, corresponding to disaccharide structural elements present in bacterial polysaccharides. Stereochemical differences at or adjacent to the glycosidic linkage were explored by solution state NMR spectroscopy using one-dimensional 1H,1H-NOESY NMR experiments to obtain transglycosidic proton-proton distances and one- and two-dimensional heteronuclear NMR experiments to obtain 3JCH transglycosidic coupling constants related to torsion angles phi and psi. Computed effective proton-proton distances from molecular dynamics (MD) simulations showed excellent agreement to experimentally derived distances for the alpha-(1 -> 3)-linked disaccharides and revealed that for the bimodal distribution at the psi torsion angle for the alpha-(1 -> 4)-linked disaccharide experiment and simulation were at variance with each other, calling for further force field developments. The MD simulations disclosed a highly intricate inter-residue hydrogen bonding pattern for the alpha-(1 -> 4)-linked disaccharide, including a nonconventional hydrogen bond between H5 ' in the glucosyl residue and O3 in the galactosyl residue, supported by a large downfield 1H NMR chemical shift displacement compared to alpha-d-Glcp-OMe. Comparison of population distributions of the glycosidic torsion angles phi and psi in the disaccharide entities to those of corresponding crystal structures highlighted the potential importance of solvation on the preferred conformation. The importance of solvation on the preferred conformation of saccharides in solution and in crystals is unraveled by solution-state NMR and computational MD studies of solvated disaccharides. Crystal structures containing solvated glycan structures have glycosidic linkage conformations similar to those of the carbohydrate molecules in solution. 

Keywords
Dynamics, Glycan, Hydrogen bond, NOESY, PDB
National Category
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-226551 (URN)10.1002/chem.202304047 (DOI)001153358500001 ()38180821 (PubMedID)2-s2.0-85182823197 (Scopus ID)
Available from: 2024-02-15 Created: 2024-02-15 Last updated: 2024-02-15
Widmalm, G. (2024). Glycan Shape, Motions, and Interactions Explored by NMR Spectroscopy. JACS Au, 4(1), 20-39
Open this publication in new window or tab >>Glycan Shape, Motions, and Interactions Explored by NMR Spectroscopy
2024 (English)In: JACS Au, E-ISSN 2691-3704, Vol. 4, no 1, p. 20-39Article, review/survey (Refereed) Published
Abstract [en]

Glycans in the form of oligosaccharides, polysaccharides, and glycoconjugates are ubiquitous in nature, and their structures range from linear assemblies to highly branched and decorated constructs. Solution state NMR spectroscopy facilitates elucidation of preferred conformations and shapes of the saccharides, motions, and dynamic aspects related to processes over time as well as the study of transient interactions with proteins. Identification of intermolecular networks at the atomic level of detail in recognition events by carbohydrate-binding proteins known as lectins, unraveling interactions with antibodies, and revealing substrate scope and action of glycosyl transferases employed for synthesis of oligo- and polysaccharides may efficiently be analyzed by NMR spectroscopy. By utilizing NMR active nuclei present in glycans and derivatives thereof, including isotopically enriched compounds, highly detailed information can be obtained by the experiments. Subsequent analysis may be aided by quantum chemical calculations of NMR parameters, machine learning-based methodologies and artificial intelligence. Interpretation of the results from NMR experiments can be complemented by extensive molecular dynamics simulations to obtain three-dimensional dynamic models, thereby clarifying molecular recognition processes involving the glycans.

Keywords
carbohydrate, conformation, coupling constant, molecular dynamics, recognition, relaxation, simulation
National Category
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-225978 (URN)10.1021/jacsau.3c00639 (DOI)001146747200001 ()38274261 (PubMedID)2-s2.0-85182010162 (Scopus ID)
Available from: 2024-01-31 Created: 2024-01-31 Last updated: 2024-01-31Bibliographically approved
Dorst, K., Engström, O., Angles d'Ortoli, T., Mobarak, H., Ebrahemi, A., Fagerberg, U., . . . Widmalm, G. (2024). On the influence of solvent on the stereoselectivity of glycosylation reactions. Carbohydrate Research, 535, Article ID 109010.
Open this publication in new window or tab >>On the influence of solvent on the stereoselectivity of glycosylation reactions
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2024 (English)In: Carbohydrate Research, ISSN 0008-6215, E-ISSN 1873-426X, Vol. 535, article id 109010Article in journal (Refereed) Published
Abstract [en]

Methodology development in carbohydrate chemistry entails the stereoselective formation of C-O bonds as a key step in the synthesis of oligo- and polysaccharides. The anomeric selectivity of a glycosylation reaction is affected by a multitude of parameters, such as the nature of the donor and acceptor, activator/promotor system, temperature and solvent. The influence of different solvents on the stereoselective outcome of glycosylation reactions employing thioglucopyranosides as glycosyl donors with a non-participating protecting group at position 2 has been studied. A large change in selectivity as a function of solvent was observed and a correlation between selectivity and the Kamlet-Taft solvent parameter pi* was found. Furthermore, molecular modeling using density functional theory methodology was conducted to decipher the role of the solvent and possible reaction pathways were investigated.

Keywords
DFT, Donor, Glucopyranoside, Promotor, Thioglycoside
National Category
Biocatalysis and Enzyme Technology
Identifiers
urn:nbn:se:su:diva-226576 (URN)10.1016/j.carres.2023.109010 (DOI)001155958200001 ()38181544 (PubMedID)2-s2.0-85181825919 (Scopus ID)
Available from: 2024-02-14 Created: 2024-02-14 Last updated: 2024-02-14Bibliographically approved
Wiseman, B., Widmalm, G. & Högbom, M. (2023). Alternating L4 loop architecture of the bacterial polysaccharide co-polymerase WzzE. Communications Biology, 6(1), Article ID 802.
Open this publication in new window or tab >>Alternating L4 loop architecture of the bacterial polysaccharide co-polymerase WzzE
2023 (English)In: Communications Biology, E-ISSN 2399-3642, Vol. 6, no 1, article id 802Article in journal (Refereed) Published
Abstract [en]

Lipopolysaccharides such as the enterobacterial common antigen are important components of the enterobacterial cell envelope that act as a protective barrier against the environment and are often polymerized by the inner membrane bound Wzy-dependent pathway. By employing cryo-electron microscopy we show that WzzE, the co-polymerase component of this pathway that is responsible for the length modulation of the enterobacterial common antigen, is octameric with alternating up-down conformations of its L4 loops. The alternating up-down nature of these essential loops, located at the top of the periplasmic bell, are modulated by clashing helical faces between adjacent protomers that flank the L4 loops around the octameric periplasmic bell. This alternating arrangement and a highly negatively charged binding face create a dynamic environment in which the polysaccharide chain is extended, and suggest a ratchet-type mechanism for polysaccharide elongation. Cryo-EM structure of bacterial polysaccharide co-polymerase WzzE provides insight into possible mechanisms of lipopolysaccharide elongation and length regulation.

National Category
Other Biological Topics Other Chemistry Topics
Identifiers
urn:nbn:se:su:diva-220901 (URN)10.1038/s42003-023-05157-7 (DOI)001048682700004 ()37532793 (PubMedID)2-s2.0-85166437281 (Scopus ID)
Available from: 2023-09-18 Created: 2023-09-18 Last updated: 2023-09-18Bibliographically approved
Plazinski, W., Angles d'Ortoli, T. & Widmalm, G. (2023). Conformational flexibility of the disaccharide & beta;-l-Fucp-(1 & RARR;4)-& alpha;-d-Glcp-OMe as deduced from NMR spectroscopy experiments and computer simulations. Organic and biomolecular chemistry (34)
Open this publication in new window or tab >>Conformational flexibility of the disaccharide & beta;-l-Fucp-(1 & RARR;4)-& alpha;-d-Glcp-OMe as deduced from NMR spectroscopy experiments and computer simulations
2023 (English)In: Organic and biomolecular chemistry, ISSN 1477-0520, E-ISSN 1477-0539, no 34Article in journal (Refereed) Published
Abstract [en]

Carbohydrates in biological systems are referred to as glycans and modification of their structures is a hallmark indicator of disease. Analysis of the three-dimensional structure forms the basis for further insight into how they function and comparison of crystal structure with solution-state conformation(s) is particularly relevant, which has been performed for the disaccharide & beta;-l-Fucp-(1 & RARR;4)-& alpha;-d-Glcp-OMe. In water solution the conformational space at the glycosidic linkage between the two sugar residues is identified from molecular dynamics (MD) simulations as having a low-energy exo-syn conformation, deviating somewhat from the solid-state conformation, and two anti-conformational states, i.e., anti-& phi; and anti-& psi;, indicating flexibility at the glycosidic linkage. NMR data were obtained from 1D H-1,H-1-NOESY and STEP-NOESY experiments, measurement of transglycosidic (3)J(CH) coupling constants and NMR spin-simulation. The free energy profile of the & omega; torsion angle computed from MD simulation was in excellent agreement with the rotamer distribution from NMR experiment being for gt:gg:tg 38 : 53 : 9, respectively, with a proposed inter-residue O5 & PRIME;MIDLINE HORIZONTAL ELLIPSISHO6 hydrogen bond being predominant in the gg rotamer. Quantum mechanics methodology was used to calculate transglycosidic NMR (3)J(CH) coupling constants, averaged over a conformational ensemble of structures representing various rotamers of exocyclic groups, in good to excellent agreement with Karplus-type relationships previously developed. Furthermore, H-1 and C-13 NMR chemical shifts were calculated using the same methodology and were found to be in excellent agreement with experimental data.

National Category
Chemical Sciences
Identifiers
urn:nbn:se:su:diva-221324 (URN)10.1039/d3ob01153d (DOI)001048518400001 ()37584331 (PubMedID)2-s2.0-85169173471 (Scopus ID)
Available from: 2023-09-19 Created: 2023-09-19 Last updated: 2023-09-19Bibliographically approved
Eriksson, L. & Widmalm, G. (2023). Crystal Structure of Methyl 3-O-α-D-Glucopyranosyl 2-Acetamido-2-Deoxy-α-D-Galactopyranoside Hydrate. Journal of Chemical Crystallography, 53(3), 400-406
Open this publication in new window or tab >>Crystal Structure of Methyl 3-O-α-D-Glucopyranosyl 2-Acetamido-2-Deoxy-α-D-Galactopyranoside Hydrate
2023 (English)In: Journal of Chemical Crystallography, ISSN 1074-1542, E-ISSN 1572-8854, Vol. 53, no 3, p. 400-406Article in journal (Refereed) Published
Abstract [en]

Methyl 3-O-α-D-glucopyranosyl 2-acetamido-2-deoxy-α-D-galactopyranoside as a monohydrate, C15H27NO11·H2O, crystallizes in space group P212121, with four molecules in the unit cell. It constitutes the methyl glycoside of the carbohydrate part of the teichoic acid type polysaccharide from Micrococcus sp. A1, in which the disaccharides are joined through phosphodiester linkages. The conformation of the disaccharide is described by the glycosidic torsion angles ϕ =  − 31° and ψ =  + 1°, and the hydroxymethyl groups of the constituent monosaccharides are present in the gg and gt conformations for the sugar residues having the gluco- and galacto-configuration, respectively. For the N-acetyl group at C2 of the galactosamine residue the torsion angle τH = 147°, i.e., the amide proton has an antiperiplanar relationship to H2 of the sugar ring. The structure shows extensive hydrogen bonding along the a-direction, including the water molecule, and forms sheets with hydrophilic interactions within the sheets as a result of hydrogen bonding between disaccharides as well as hydrophobic interactions between the sheets, in particular, amongst methyl groups of the N-acetyl group of the α-D-GalpNAc residue in the disaccharides.

Keywords
Crystal structure, Carbohydrate, Disaccharide, Hydrogen bonding, DFT
National Category
Other Chemistry Topics
Identifiers
urn:nbn:se:su:diva-216325 (URN)10.1007/s10870-023-00979-z (DOI)000947447600001 ()2-s2.0-85149721653 (Scopus ID)
Available from: 2023-04-11 Created: 2023-04-11 Last updated: 2023-10-04Bibliographically approved
Kwon, J., Ruda, A., Azurmendi, H. F., Zarb, J., Battistel, M. D., Liao, L., . . . Freedberg, D. I. (2023). Glycan Stability and Flexibility: Thermodynamic and Kinetic Characterization of Nonconventional Hydrogen Bonding in Lewis Antigens. Journal of the American Chemical Society, 145(18), 10022-10034
Open this publication in new window or tab >>Glycan Stability and Flexibility: Thermodynamic and Kinetic Characterization of Nonconventional Hydrogen Bonding in Lewis Antigens
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2023 (English)In: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 145, no 18, p. 10022-10034Article in journal (Refereed) Published
Abstract [en]

We provide evidence for CH-based nonconventional hydrogen bonds (H-bonds) for 10 Lewis antigens and two of their rhamnose analogues. We also characterize the thermodynamics and kinetics of the H-bonds in these molecules and present a plausible explanation for the presence of nonconventional H-bonds in Lewis antigens. Using an alternative method to simultaneously fit a series of temperature-dependent fast exchange nuclear magnetic resonance (NMR) spectra, we determined that the H-bonded conformation is favored by ∼1 kcal/mol over the non-H-bonded conformation. Additionally, a comparison of temperature-dependent 13C linewidths in various Lewis antigens and the two rhamnose analogues reveals H-bonds between the carbonyl oxygen of the N-acetyl group of N-acetylglucosamine and the OH2 group of galactose/fucose. The data presented herein provide insight into the contribution of nonconventional H-bonding to molecular structure and could therefore be used for the rational design of therapeutics.

National Category
Chemical Sciences
Identifiers
urn:nbn:se:su:diva-217353 (URN)10.1021/jacs.2c13104 (DOI)000981760800001 ()37099481 (PubMedID)2-s2.0-85156142488 (Scopus ID)
Available from: 2023-05-29 Created: 2023-05-29 Last updated: 2023-05-29Bibliographically approved
Ruda, A., Aytenfisu, A. H., Angles d'Ortoli, T., MacKerell Jr, A. D. & Widmalm, G. (2023). Glycosidic α-linked mannopyranose disaccharides: an NMR spectroscopy and molecular dynamics simulation study employing additive and Drude polarizable force fields. Physical Chemistry, Chemical Physics - PCCP, 25(4), 3042-3060
Open this publication in new window or tab >>Glycosidic α-linked mannopyranose disaccharides: an NMR spectroscopy and molecular dynamics simulation study employing additive and Drude polarizable force fields
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2023 (English)In: Physical Chemistry, Chemical Physics - PCCP, ISSN 1463-9076, E-ISSN 1463-9084, Vol. 25, no 4, p. 3042-3060Article in journal (Refereed) Published
Abstract [en]

D-Mannose is a structural component in N-linked glycoproteins from viruses and mammals as well as in polysaccharides from fungi and bacteria. Structural components often consist of D-Manp residues joined via α-(1→2)-, α-(1→3)-, α-(1→4)- or α-(1→6)-linkages. As models for these oligo- and polysaccharides, a series of mannose-containing disaccharides have been investigated with respect to conformation and dynamics. Translational diffusion NMR experiments were performed to deduce rotational correlation times for the molecules, 1D 1H,1H-NOESY and 1D 1H,1H-T-ROESY NMR experiments were carried out to obtain inter-residue proton–proton distances and one-dimensional long-range and 2D J-HMBC experiments were acquired to gain information about conformationally dependent heteronuclear coupling constants across glycosidic linkages. To attain further spectroscopic data, the doubly 13C-isotope labeled α-D-[1,2-13C2]Manp-(1→4)-α-D-Manp-OMe was synthesized thereby facilitating conformational analysis based on 13C,13C coupling constants as interpreted by Karplus-type relationships. Molecular dynamics simulations were carried out for the disaccharides with explicit water as solvent using the additive CHARMM36 and Drude polarizable force fields for carbohydrates, where the latter showed broader population distributions. Both simulations sampled conformational space in such a way that inter-glycosidic proton–proton distances were very well described whereas in some cases deviations were observed between calculated conformationally dependent NMR scalar coupling constants and those determined from experiment, with closely similar root-mean-square differences for the two force fields. However, analyses of dipole moments and radial distribution functions with water of the hydroxyl groups indicate differences in the underlying physical forces dictating the wider conformational sampling with the Drude polarizable versus additive C36 force field and indicate the improved utility of the Drude polarizable model in investigating complex carbohydrates.

National Category
Chemical Sciences
Identifiers
urn:nbn:se:su:diva-214350 (URN)10.1039/d2cp05203b (DOI)000907979300001 ()36607620 (PubMedID)2-s2.0-85146192918 (Scopus ID)
Available from: 2023-02-02 Created: 2023-02-02 Last updated: 2023-02-02Bibliographically approved
Gao, Y., Widmalm, G. & Im, W. (2023). Modeling and Simulation of Bacterial Outer Membranes with Lipopolysaccharides and Capsular Polysaccharides. Journal of Chemical Information and Modeling, 63(5), 1592-1601
Open this publication in new window or tab >>Modeling and Simulation of Bacterial Outer Membranes with Lipopolysaccharides and Capsular Polysaccharides
2023 (English)In: Journal of Chemical Information and Modeling, ISSN 1549-9596, E-ISSN 1549-960X, Vol. 63, no 5, p. 1592-1601Article in journal (Refereed) Published
Abstract [en]

Capsule is one of the common virulence factors in Gram-negative bacteria protecting pathogens from host defenses and consists of long-chain capsular polysaccharides (CPS) anchored in the outer membrane (OM). Elucidating structural properties of CPS is important to understand its biological functions as well as the OM properties. However, the outer leaflet of the OM in current simulation studies is represented exclusively by LPS due to the complexity and diversity of CPS. In this work, representative Escherichia coli CPS, KLPS (a lipid A-linked form) and KPG (a phosphatidylglycerol-linked form), are modeled and incorporated into various symmetric bilayers with co-existing LPS in different ratios. All-atom molecular dynamics simulations of these systems have been conducted to characterize various bilayer properties. Incorporation of KLPS makes the acyl chains of LPS more rigid and ordered, while incorporation of KPG makes them less ordered and flexible. These results are consistent with the calculated area per lipid (APL) of LPS, in which the APL of LPS becomes smaller when KLPS is incorporated, whereas it gets larger when KPG is included. Torsional analysis reveals that the influence of the CPS presence on the conformational distributions of the glycosidic linkages of LPS is small, and minor differences are also detected for the inner and outer regions of the CPS. Combined with previously modeled enterobacterial common antigens (ECAs) in the form of mixed bilayers, this work provides more realistic OM models as well as the basis for characterization of interactions between the OM and OM proteins.

National Category
Chemical Sciences Biological Sciences
Identifiers
urn:nbn:se:su:diva-216436 (URN)10.1021/acs.jcim.3c00072 (DOI)000951958600001 ()36802606 (PubMedID)2-s2.0-85149337040 (Scopus ID)
Available from: 2023-05-05 Created: 2023-05-05 Last updated: 2023-05-08Bibliographically approved
Dorst, K. & Widmalm, G. (2023). NMR chemical shift prediction and structural elucidation of linker-containing oligo- and polysaccharides using the computer program CASPER. Carbohydrate Research, 533, Article ID 108937.
Open this publication in new window or tab >>NMR chemical shift prediction and structural elucidation of linker-containing oligo- and polysaccharides using the computer program CASPER
2023 (English)In: Carbohydrate Research, ISSN 0008-6215, E-ISSN 1873-426X, Vol. 533, article id 108937Article in journal (Refereed) Published
Abstract [en]

Carbohydrate structures containing alkyl groups as aglycones are useful for investigating enzyme activity and glycan-protein interactions. Moreover, linker-containing oligosaccharides with a spacer group are commonly used to print glycan microarrays or to prepare protein-conjugates as vaccine candidates. The structural accuracy of these synthesized glycans are essential for interpretation of results from biological experiments in which the compounds have been used and NMR spectroscopy can unravel and confirm their structures. An approach for efficient 1H and 13C NMR chemical shift assignments employed a parallel NOAH-10 measurement followed by NMR spin-simulation to refine the 1H NMR chemical shifts, as exemplified for a disaccharide with an azidoethyl group as an aglycone, the NMR chemical shifts of which have been used to enhance the quality of CASPER (http://www.casper.organ.su.se/casper/). The CASPER program has been further developed to aid characterization of linker-containing oligo- and polysaccharides, either by chemical shift prediction for comparison to experimental NMR data or as structural investigation of synthesized glycans based on acquired unassigned NMR data. The ability of CASPER to elucidate structures of linker-containing oligosaccharides is demonstrated and comparisons to assigned or unassigned NMR data show the utility of CASPER in supporting a proposed oligosaccharide structure. Prediction of NMR chemical shifts of an oligosaccharide, corresponding to the repeating unit of an O-antigen polysaccharide, having a linker as an aglycone and a non-natural substituent derivative thereof are presented to exemplify the diversity of structures handled. Furthermore, NMR chemical shift predictions of synthesized polysaccharides, corresponding to bacterial polysaccharides, containing a linker are described showing that in addition to oligosaccharide structures also polysaccharide structures having an aglycone spacer group can be analyzed by CASPER.

National Category
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-223149 (URN)10.1016/j.carres.2023.108937 (DOI)37734222 (PubMedID)2-s2.0-85171632396 (Scopus ID)
Funder
Swedish Research Council, 2022-03014Knut and Alice Wallenberg Foundation
Available from: 2023-10-23 Created: 2023-10-23 Last updated: 2023-12-19Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0001-8303-4481

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