Inter‐residual Hydrogen Bonding in Carbohydrates Unraveled by NMR Spectroscopy and Molecular Dynamics Simulations

Abstract

Carbohydrates, also known as glycans in biological systems, are omnipresent in nature where they as glycoconjugates occur as oligo‐ and polysaccharides linked to lipids and proteins. Their three‐dimensional structure is defined by two or three torsion angles at each glycosidic linkage. In addition, transglycosidic hydrogen bonding between sugar residues may be important. Herein we investigate the presence of these inter‐residue interactions by NMR spectroscopy in D2O/[D6]DMSO (70:30) or D2O and by molecular dynamics (MD) simulations with explicit water as solvent for disaccharides with structural elements α‐d‐Manp‐(1→2)‐d‐Manp, β‐d‐GlcpNAc‐(1→2)‐d‐Manp, and α‐d‐Glcp‐(1→4)‐β‐d‐Glcp, all of which have been suggested to exhibit inter‐residue hydrogen bonding. For the disaccharide β‐d‐GlcpNAc‐(1→2)‐β‐d‐Manp‐OMe, the large extent of O5′⋅⋅⋅HO3 hydrogen bonding as seen from the MD simulation is implicitly supported by the 1H NMR chemical shift and 3JHO3,H3 value of the hydroxy proton. In the case of α‐d‐Glcp‐(1→4)‐β‐d‐Glcp‐OMe, the existence of a transglycosidic hydrogen bond O2′⋅⋅⋅HO3 was proven by the presence of a cross‐peak in 1H,13C HSQC‐TOCSY experiments as a result of direct TOCSY transfer between HO3 of the reducing end residue and H2′ (detected at C2′) of the terminal residue. The occurrence of inter‐residue hydrogen bonding, albeit transient, is judged important for the stabilization of three‐dimensional structures, which may be essential in maintaining a conformational state for carbohydrate–protein interactions of glycans to take place in biologically important environments.

Pyranose Ring Puckering Thermodynamics for Glycan Monosaccharides Associated with Vertebrate Proteins

“…In addition to demonstrating that the CHARMM force field reliably models ring puckering across this set diverse of molecules, the results show that doing so is possible with a single set of self-consistent force-field parameters developed using a standardized force field parametrization protocol [ 36 , 38 ]. This, in combination with examples of CHARMM force field studies on glycosidic linkages [ 91 , 92 , 93 , 94 , 95 , 96 ], lends confidence to the application of these parameters in the modeling of carbohydrate-containing protein systems, such as glycoproteins and proteoglycans as well as transmembrane proteins in glycolipid-containing bilayers. Accurate simulations for these types of systems can help expand the frontiers of protein structural biology by bridging gaps in experimental approaches for characterizing carbohydrate-containing protein systems.…”

Helical polysaccharides

“…However, the NOE short range (up to 4 Å) and large timescale (50 ms to 1 second) often produced uncertain results . Other methods, such as measurement of residual dipolar coupling (RDC), analyses of J ‐couplings across O ‐glycosidic linkages, and hydrogen bond studies are useful and more reliable tools . In contrast to protein, the use of NMR suffers from the chemical shift degeneracy and intrinsic flexibility of polysaccharides.…”

Selectively Deoxyfluorinated N‐Acetyllactosamine Analogues as 19F NMR Probes to Study Carbohydrate‐Galectin Interactions

“…Conversely, deoxyfluorination at the C3 of the glucosamine unit gave the lowest value of log P in the series. We speculate that this high hydrophilicity of the 3F‐LacNAc analogue may result from disruption of the transient inter‐residual hydrogen bond between O3‐H and the galactose ring oxygen O5′ (O5′⋅⋅⋅H−O3) upon deoxyfluorination [47] . The hydrogen‐bonded O3‐H hydroxyl is probably less solvated by water molecules than are other hydroxyls, because its hydrogen‐bonding capability towards water is reduced, [48] and its contribution to hydrophilicity of LacNAc is therefore less significant.…”

“…We speculate that this high hydrophilicity of the 3F-LacNAc analogue may result from disruption of the transient interresidual hydrogen bond between O3-H and the galactose ring oxygen O5' (O5'•••HÀ O3) upon deoxyfluorination. [47] The hydrogen-bonded O3-H hydroxyl is probably less solvated by water molecules than are other hydroxyls, because its hydrogenbonding capability towards water is reduced, [48] and its contribution to hydrophilicity of LacNAc is therefore less significant. Accordingly, deoxyfluorination of O3 hydroxyl to 3F-LacNAc 6 will not increase lipophilicity to such extent as deoxyfluorination at other positions, also because it will expose the ring oxygen O5' to increased solvation by water.…”