(E)-Alkene and Ethylene Isosteres Substantially Alter the Hydrogen-Bonding Network in Class II MHC A^q/Glycopeptide Complexes and Affect T-Cell Recognition

Abstract

The structural basis for antigen presentation by class II major histocompatibility complex (MHC) proteins to CD4(+) T-cells is important for understanding and possibly treating autoimmune diseases. In the work described in this paper, (E)-alkene and ethylene amide-bond isosteres were used to investigate the effect of removing hydrogen-bonding possibilities from the CII259-270 glycopeptide, which is bound by the arthritis-associated murine A(q) class II MHC protein. The isostere-modified glycopeptides showed varying and unexpectedly large losses of A(q) binding that could be linked to the dynamics of the system. Molecular dynamics (MD) simulations revealed that the backbone of CII259-270 and the A(q) protein are able to form up to 11 hydrogen bonds, but fewer than this number are present at any one time. Most of the strong hydrogen-bond interactions were formed by the N-terminal part of the glycopeptide, i.e., in the region where the isosteric replacements were made. The structural dynamics also revealed that hydrogen bonds were strongly coupled to each other; the loss of one hydrogen-bond interaction had a profound effect on the entire hydrogen-bonding network. The A(q) binding data revealed that an ethylene isostere glycopeptide unexpectedly bound more strongly to A(q) than the corresponding (E)-alkene, which is in contrast to the trend observed for the other isosteres. Analysis of the MD trajectories revealed that the complex conformation of this ethylene isostere was structurally different and had an altered molecular interaction pattern compared to the other A(q)/glycopeptide complexes. The introduced amide-bond isosteres also affected the interactions of the glycopeptide/A(q) complexes with T-cell receptors. The dynamic variation of the patterns and strengths of the hydrogen-bond interactions in the class II MHC system is of critical importance for the class II MHC/peptide/TCR signaling system.

Hydroxyethylene isosteres introduced in type II collagen fragments substantially alter the structure and dynamics of class II MHC Aq/glycopeptide complexes

“…At this position the corresponding ketomethylene-isostere 4 is the only modified glycopeptide so far that has been shown to bind equally well to A q as 1. [12][13][14] The T-cell response was investigated in a cell-based assay, where IL-2 secretion of A q restricted T-cell hybridoma lines was measured after incubation with antigen-presenting spleen cells expressing A q and increasing concentrations of the glycopeptides 1, 2 or 3 (Fig. 4).…”

“…That HCQ.10 and 22a1-7e were more sensitive towards the introduced modifications in the backbone compared to HCQ.3 and HMIR.2 has also been seen in previous studies. 13,14 Analysis of glycopeptide/A q -complexes by MD simulations Glycopeptides 1-4 were modelled into A q to understand if the structures of the complexes correlated with the binding affinity for A q and with the subsequent T cell responses. We found that the modified glycopeptides 2-4 displayed a binding mode almost identical to that of the native glycopeptide 1 in their respective complexes.…”

“…6 The minimal epitope in 1 required for binding to A q , and stimulation of CII restricted T cell hybridomas obtained in CIA, consists of amino acid residues Ile 260 -Gln 267 . 7 Antigenic peptides bind between the α 1 -and β 1 -helices of class II MHC proteins † Electronic supplementary information (ESI) available: 1 H NMR and 13 C NMR spectra of all new isolated compounds, RMSD for A q vs. simulation time, RMSD for the peptide binding groove vs. simulation time, RMSD for glycopeptides 1-4 vs. simulation time, RMSD for Ile 260 -Glu 266 sequences vs. simulation time, RMSD for the α 1 helix vs. simulation time, RMSD for the β 1 helix vs. simulation time, RMSD for the β-sheet of the α 1 β 1 domain vs. simulation time, the number of frames occupying each cluster for the MD simulations, superposition of frames from the cluster analysis displaying the glycopeptide/A q complex, superposition of frames from the cluster analysis displaying the glycopeptides 1-4, visualization of hydrogen bonds formed between A q and glycopeptides 1-4. See DOI: 10.1039/c5ob00395d through a hydrogen bond network and by anchoring of sidechains in pockets in the binding groove.…”

“…11 Modification of glycopeptides, through the incorporation of amide bond isosteres, has been used to probe molecular interactions in the MHC/glycopeptide/TCR system. [12][13][14][15][16][17] For instance, aza-amino acids, 15 (E)-alkene and ethylene isosteres 13 and oxazole modifications 12 have been employed. The Ile 260 -GalHyl 264 fragment of glycopeptide 1 has been the target for the majority of these modifications as it contains essential features, such as the two MHC anchor residues 9 and the galactose moiety crucial for TCR recognition.…”

“…4,11 Incorporation of isosteres into 1 revealed the importance of having an intact network of hydrogen bonds for the stability of the complex with A q . 13 Furthermore, the dynamics of the complex were found to govern the response from the TCR; complexes that were more rigid than those of A q and 1 resulted in weaker T cell responses. 10,13 In this study, we further explore the molecular interactions in the MHC/glycopeptide/TCR system that are crucial for the development of CIA, and most likely also for RA, by synthesis and evaluation of two additional amide bond isosteres incorporated into glycopeptide 1.…”