Molecular Recognition of β‐O‐GlcNAc Glycopeptides by a Lectin‐Like Receptor: Binding Modulation by the Underlying Ser or Thr Amino Acids

Abstract

The binding properties of different carbohydrates and glycopeptides containing the β-O-2-deoxy-2-(N-acetyl)-D-glucosaminyl (β-O-GlcNAc) to a synthetically prepared lectin-like receptor have been analyzed. The study combines the use of NMR spectroscopy experiments with extensive MD simulations in explicit water. Notably, the presence of a key hydrogen bond between the receptor and the OMe group of the β-O-GlcNAc-OMe derivative appears to be responsible for the high selectivity observed for this compound. In addition, to study the effect on the binding of the underlying amino acid, we have prepared different model glycopeptides, which include the non-natural α-methylserine and α-methylthreonine as underlying amino acids. Interestingly, the presence of a methyl group decreases the affinity constant, especially in those cases in which a β-methyl group is present. As a result, the serine-containing glycopeptide exhibited the highest affinity constant of the glycopeptides, and the threonine derivative showed the lowest one. This low selectivity could have its origin in the difficulty to form both specific hydrogen bonds and hydrophobic (CH-π) contacts.

OGlcNAcylation and Phosphorylation Have Similar Structural Effects in α-Helices: Post-Translational Modifications as Inducible Start and Stop Signals in α-Helices, with Greater Structural Effects on Threonine Modification

“…28 In addition, Corzana et al have observed greater conformational restriction in ThrOGlcNAc over SerOGlcNAc amino acids, as well as more generally in glycosylated threonine over glycosylated serine amino acids. 28,42,125−127 The data herein are consistent with all of these observations, with structural effects of threonine phosphorylation and threonine OGlcNAcylation observed herein greater than those of the same post-translational modifications on serine. In particular, the significant conformational restriction at phosphothreonine, combined with larger induced structural effects seen by CD and by NMR, suggests a special role for phosphothreonine residues in protein structure.…”

Advances in chemical probing of protein O-GlcNAc glycosylation: structural role and molecular mechanisms

“… 89 Using 1 H NMR titrations in aqueous solution, this biphenyl-based receptor was found to bind preferentially β-GlcNAc-OMe (association constant, K a = 630 M −1 ) compared to other monosaccharides ( e.g. glucose, GalNAc, Neu5Ac) and bound even better an O -GlcNAcylated CKII model decapeptide ( K a = 1000 M −1 , confirmed by induced CD), due to further backbone contacts and an enhancement of a key hydrogen bond interaction between the glycopeptide and the receptor, as subsequently observed by MD simulations by Corzana et al 251 Despite having similar affinities, this synthetic receptor outperforms the natural WGA lectin particularly in terms of selectivity, being able to discriminate between O -GlcNAc and di- N -acetyl chitobiose and an Asn-linked GlcNAc derivative (estimated K a ≈ 4 M −1 ). Bidimensional NMR experiments including NOESY data integrated as experimental constraints in molecular computational calculations revealed the expected structure for the β-GlcNAc-OMe/receptor complex ( Fig.…”

“…Based on the promising features and structural information of this β- O -GlcNAc synthetic receptor, Corzana et al expanded the molecular recognition study to probe the binding properties of the simplest Ser/Thr-containing β- O -GlcNAc system by using analogous NMR experiments in combination with MD simulations. 251 Moreover, to explore the effect of the underlying amino acid on glycopeptide conformation and binding, the related O -GlcNAc derivatives incorporating unnatural α-methylserine (MeSer) and α-methylthreonine (MeThr) amino acids, previously shown to stabilise folded conformations in model β- O -glucopeptides, 252 were also investigated. Concerning the free state conformational analysis, while the peptide backbone of both unnatural glycopeptides adopted similar, mainly helix-like conformations attributed to the α,α-disubstituted amino acid incorporation, the MeThr analogue was more rigid displaying an eclipsed conformation for the glycosidic linkage due to the β-methyl group, as previously observed for the corresponding β- O -GlcNAc-Ser/Thr model compounds.…”

Cyclohexane Ring as a Tool to Select the Presentation of the Carbohydrate Moiety in Glycosyl Amino Acids

“…In this regard, we have recently reported the different sugar presentation showed by Ser and Thr glycopeptides [3] and its influence on the binding to a synthetically prepared lectin-like receptor. [4] In addition, it is necessary to study in detail the interactions between the sugar and the underlying amino acid because they play a crucial role in the presentation of the carbohydrate moiety and, consequently, in the interactions with the corresponding biological receptors.…”