Dynamics of exocyclic groups in the Escherichia coli O91 O-antigen polysaccharide in solution studied by carbon-13 NMR relaxation

Šoltésová, Mária; Kowalewski, Józef; Widmalm, Göran

doi:10.1007/s10858-013-9763-5

Cited by 6 publications

(8 citation statements)

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“…18 By utilizing the site-specifically labeled materials it was possible to determine dynamics of the constituent sugars in the PS as well as the dynamics of the exocyclic hydroxymethyl groups of three of the sugars of the RUs. 19,20 Also, NMR data facilitated the determination of the chain length of the polymer to be ~10 repeating units, which was consistent with a narrow distribution deduced from a MALDI-TOF mass spectrometry experiment.…”

Section: Introductionsupporting

confidence: 74%

“…18 In order to be able to easily refer to aspects and results from the previous studies we continue to use to the original designation of the sugar residues denoted by A -E (Figure 1). The 1 H and 13 C NMR chemical shift assignments at 338 K were thus available from the previous study and minor chemical shift displacements besides those already documented in 13 C NMR relaxation studies at 310 K for [1-13 C]-as well as [6-13 C]-labeled E. coli O91 PS 19,20 were readily accounted for by standard 2D NMR experiments. The NMR chemical shift assignments of H6pro-R and H6pro-S of the hydroxymethyl group in the galactose residue (A) were obtained by a 1 H, 1 H-NOESY NMR experiment 67 considering the NOEs between H4 and the two hydroxymethyl group protons in conjunction with the anticipated rotameric distribution at the ω torsion angle of galactose.…”

Section: Nmr Experimentsmentioning

confidence: 99%

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Conformational Dynamics of the Lipopolysaccharide from Escherichia coli O91 Revealed by Nuclear Magnetic Resonance Spectroscopy and Molecular Simulations

et al. 2017

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The outer leaflet of the outer membrane in Gram-negative bacteria contains lipopolysaccharides (LPS) as a major component, and the outer membrane provides a physical barrier and protection against hostile environments. The enterohemorrhagic Escherichia coli of serogroup O91 has an O-antigen polysaccharide (PS) with five sugar residues in the repeating unit (RU), and the herein studied O-antigen PS contains ∼10 RUs. H-C HSQC-NOESY experiments on a 1-C-labeled PS were employed to deduce H-H cross-relaxation rates and transglycosidic J related to the ψ torsional angles were obtained by H-H NOESY experiments. Dynamical parameters were calculated from the molecular dynamics (MD) simulations of the PS in solution and compared to those from C nuclear magnetic resonance (NMR) relaxation studies. Importantly, the MD simulations can reproduce the dynamical behavior of internal correlation times along the PS chain. Two-dimensional free energy surfaces of glycosidic torsion angles delineate the conformational space available to the O-antigen. Although similar with respect to populated states in solution, the O-antigen in LPS bilayers has more extended chains as a result of spatial limitations due to close packing. Calcium ions are highly abundant in the phosphate-containing core region mediating LPS-LPS association that is crucial for maintaining bilayer integrity, and the negatively charged O-antigen promotes a high concentration of counterbalancing potassium ions. The ensemble of structures present for the PS in solution is captured by the NMR experiments, and the similarities between the O-antigen on its own and as a constituent of the full LPS in a bilayer environment make it possible to realistically describe the LPS conformation and dynamics from the MD simulations.

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Section: Introductionsupporting

confidence: 74%

Section: Nmr Experimentsmentioning

confidence: 99%

Conformational Dynamics of the Lipopolysaccharide from Escherichia coli O91 Revealed by Nuclear Magnetic Resonance Spectroscopy and Molecular Simulations

et al. 2017

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“…Contrary to proteins, polysaccharides are not globular but extended or random structures in which different segments may display different flexibility (Martin-Pastor and Bush 2000;Soltesova et al 2013). To evaluate the effect of the temperature and the spectrometer magnetic field strength on the relaxation parameters of this kind of structures, a polysaccharide model of approximately the same molecular weight as the O-antigen polysaccharide of E. coli O142, but with a simplified labeling pattern, was selected: the [1-13 C]-labeled Oantigen polysaccharide of E. coli O91 (Fig.…”

Section: Resultsmentioning

confidence: 99%

NMR structure analysis of uniformly 13C-labeled carbohydrates

2014

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In this study, a set of nuclear magnetic resonance experiments, some of them commonly used in the study of (13)C-labeled proteins and/or nucleic acids, is applied for the structure determination of uniformly (13)C-enriched carbohydrates. Two model substances were employed: one compound of low molecular weight [(UL-(13)C)-sucrose, 342 Da] and one compound of medium molecular weight ((13)C-enriched O-antigenic polysaccharide isolated from Escherichia coli O142, ~10 kDa). The first step in this approach involves the assignment of the carbon resonances in each monosaccharide spin system using the anomeric carbon signal as the starting point. The (13)C resonances are traced using (13)C-(13)C correlations from homonuclear experiments, such as (H)CC-CT-COSY, (H)CC-NOESY, CC-CT-TOCSY and/or virtually decoupled (H)CC-TOCSY. Based on the assignment of the (13)C resonances, the (1)H chemical shifts are derived in a straightforward manner using one-bond (1)H-(13)C correlations from heteronuclear experiments (HC-CT-HSQC). In order to avoid the (1) J CC splitting of the (13)C resonances and to improve the resolution, either constant-time (CT) in the indirect dimension or virtual decoupling in the direct dimension were used. The monosaccharide sequence and linkage positions in oligosaccharides were determined using either (13)C or (1)H detected experiments, namely CC-CT-COSY, band-selective (H)CC-TOCSY, HC-CT-HSQC-NOESY or long-range HC-CT-HSQC. However, due to the short T2 relaxation time associated with larger polysaccharides, the sequential information in the O-antigen polysaccharide from E. coli O142 could only be elucidated using the (1)H-detected experiments. Exchanging protons of hydroxyl groups and N-acetyl amides in the (13)C-enriched polysaccharide were assigned by using HC-H2BC spectra. The assignment of the N-acetyl groups with (15)N at natural abundance was completed by using HN-SOFAST-HMQC, HNCA, HNCO and (13)C-detected (H)CACO spectra.

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“…Different NMR relaxation data are often measured at more than one magnetic field strength in order to obtain a sufficiently large number of observables to fit to motional parameters of dynamic models. 61,62 For small molecules in low viscosity solvents the rotational reorientation is rapid and the NMR relaxation parameters are independent of the magnetic field employed, i.e., the extreme narrowing region prevails where o 2 t c 2 { 1. 63 For a sample of [1 0 -13 C]-R2R in D 2 O at 298 K the heteronuclear NOE factor at 14.1 T was determined herein revealing NOE = 2.70, i.e., quite close to the theoretical maximum of the NOE factor, where it will be difficult to extract reliable information about internal motion from such data.…”

Section: Nmr Spectroscopy and Spin Relaxationmentioning

confidence: 99%

Flexibility at a glycosidic linkage revealed by molecular dynamics, stochastic modeling, and ¹³C NMR spin relaxation: conformational preferences of α-l-Rhap-α-(1 → 2)-α-l-Rhap-OMe in water and dimethyl sulfoxide solutions

Pendrill

Engström

Volpato

et al. 2016

Phys. Chem. Chem. Phys.

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The monosaccharide L-rhamnose is common in bacterial polysaccharides and the disaccharide α-L-Rhap-α-(1 → 2)-α-L-Rhap-OMe represents a structural model for a part of Shigella flexneri O-antigen polysaccharides. Utilization of [1'-(13)C]-site-specific labeling in the anomeric position at the glycosidic linkage between the two sugar residues facilitated the determination of transglycosidic NMR (3)JCH and (3)JCC coupling constants. Based on these spin-spin couplings the major state and the conformational distribution could be determined with respect to the ψ torsion angle, which changed between water and dimethyl sulfoxide (DMSO) as solvents, a finding mirrored by molecular dynamics (MD) simulations with explicit solvent molecules. The (13)C NMR spin relaxation parameters T1, T2, and heteronuclear NOE of the probe were measured for the disaccharide in DMSO-d6 at two magnetic field strengths, with standard deviations ≤1%. The combination of MD simulation and a stochastic description based on the diffusive chain model resulted in excellent agreement between calculated and experimentally observed (13)C relaxation parameters, with an average error of <2%. The coupling between the global reorientation of the molecule and the local motion of the spin probe is deemed essential if reproduction of NMR relaxation parameters should succeed, since decoupling of the two modes of motion results in significantly worse agreement. Calculation of (13)C relaxation parameters based on the correlation functions obtained directly from the MD simulation of the solute molecule in DMSO as solvent showed satisfactory agreement with errors on the order of 10% or less.

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Dynamics of exocyclic groups in the Escherichia coli O91 O-antigen polysaccharide in solution studied by carbon-13 NMR relaxation

Abstract: Dynamics of exocyclic groups in the Escherichia coli O91 O-antigen polysaccharide in solution studied by carbon-13 NMR relaxation. Journal of Biomolecular NMR,

Cited by 6 publications

References 40 publications

Conformational Dynamics of the Lipopolysaccharide from Escherichia coli O91 Revealed by Nuclear Magnetic Resonance Spectroscopy and Molecular Simulations

Conformational Dynamics of the Lipopolysaccharide from Escherichia coli O91 Revealed by Nuclear Magnetic Resonance Spectroscopy and Molecular Simulations

NMR structure analysis of uniformly 13C-labeled carbohydrates

Flexibility at a glycosidic linkage revealed by molecular dynamics, stochastic modeling, and ¹³C NMR spin relaxation: conformational preferences of α-l-Rhap-α-(1 → 2)-α-l-Rhap-OMe in water and dimethyl sulfoxide solutions

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Dynamics of exocyclic groups in the Escherichia coli O91 O-antigen polysaccharide in solution studied by carbon-13 NMR relaxation

Abstract: Dynamics of exocyclic groups in the Escherichia coli O91 O-antigen polysaccharide in solution studied by carbon-13 NMR relaxation. Journal of Biomolecular NMR,

Cited by 6 publications

References 40 publications

Conformational Dynamics of the Lipopolysaccharide from Escherichia coli O91 Revealed by Nuclear Magnetic Resonance Spectroscopy and Molecular Simulations

Conformational Dynamics of the Lipopolysaccharide from Escherichia coli O91 Revealed by Nuclear Magnetic Resonance Spectroscopy and Molecular Simulations

NMR structure analysis of uniformly 13C-labeled carbohydrates

Flexibility at a glycosidic linkage revealed by molecular dynamics, stochastic modeling, and 13C NMR spin relaxation: conformational preferences of α-l-Rhap-α-(1 → 2)-α-l-Rhap-OMe in water and dimethyl sulfoxide solutions

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Flexibility at a glycosidic linkage revealed by molecular dynamics, stochastic modeling, and ¹³C NMR spin relaxation: conformational preferences of α-l-Rhap-α-(1 → 2)-α-l-Rhap-OMe in water and dimethyl sulfoxide solutions