Engineering O-glycosylation points in non-extended peptides: implications for the molecular recognition of short tumor-associated glycopeptides.

The ties that bind: The incorporation of non-natural residues in the peptide backbone allows the design of O-glycosylation points in helical segments. This strategy could help to modulate the binding properties between glycopeptides and their protein receptors, such as lectins and antibodies.

The nature and sequence of the amino acid aglycone strongly modulates the conformation and dynamics effects of Tn antigen's clusters.

Synthetic oligosaccharide vaccines based on carbohydrate epitopes are currently being evaluated as potential immunotherapeutics in the treatment of cancer. In an effort to study the role that the amino acid moiety (L-serine and/or L-threonine residues) plays on the global shape of the resulting glycopeptides and on the dynamics of the carbohydrate moiety, diverse glycopeptides based on the Tn antigen have been synthesized and studied in aqueous solution by combining NMR spectroscopic experiments and molecular dynamics simulations. Our results demonstrate that although the effect of the clustering of Tn on the peptide backbone is not remarkable, it substantially modifies the dynamics, and thus, the presentation features of the carbohydrate moiety. In fact, the selected sequence has a crucial influence on both the orientation and flexibility of the sugar region. Thus, although a serine-threonine pair shows a well-defined spatial disposition of the Tn epitopes, its analogue sequence threonine-serine allows a certain degree of mobility that could favor the interaction with a diversity of receptors without a major energy penalty. These features can be explained by attending to the different conformational behavior of the glycosidic linkage of threonine-containing glycopeptides when compared with those of the serine analogues. On this basis, and taking into account that these carbohydrates interact with components of the immune system, these findings could have implications for further design of new cancer vaccines.

Serine versus threonine glycosylation: the methyl group causes a drastic alteration on the carbohydrate orientation and on the surrounding water shell.

Different behavior has been observed for the psi torsion angle of the glycosidic linkages of D-GalNAc-Ser and D-GalNAc-Thr motifs, allowing the carbohydrate moiety to adopt a completely different orientation. In addition, the fact that the water pockets found in alpha-D-GalNAc-Thr differ from those obtained for its serine analogue could be related to the different capability that the two model glycopeptides have to structure the surrounding water. This fact could have important biological inferences (i.e., antifreeze activity).