wever, although the protrusion of p1K out of the peptide-binding cleft in H-2Kb/gp33 results in a significant modification of the position of the nitrogen atom of residue p2A when compared to p1A in H2Kb/gp34 or H-2Kb/NY-gp34, this does not affect the conformation of the cluster of 1260907-17-2 web tyrosine residues that surround the Nterminal part of the peptide. Furthermore, the conformation of the stretch of peptide residues AVYNFATM bound within the peptide-binding cleft is very similar in both H2Kb/gp33 and H-2Kb/gp34. Thus although the Nand C- peptide termini play crucial roles in binding of epitopes within the peptide-binding cleft formed by the a1a2 groove, these are not always necessarily required. It has been previously demonstrated that longer peptides may protrude out of the groove and that modified and/or shorter peptides can still bind efficiently within the groove through the use of a network of water molecules within the peptide binding cleft. In the prior study by Hardy et al, T cells could be readily elicited in C57/BL6 mice following immunization with NY-gp33. MHC-I-Restricted Nitrotyrosinated Neoantigen chain of p1K from the N-terminal part of the peptide-binding cleft of H-2Kb/NY-gp33, which could potentially limit optimal recognition by the TCR. However, the question remained as to why the H-2Kb/NY-gp33 and H-2Kb/NY-gp34-specific TCR of 24H1 did not recognize H-2Kb/gp34 Nitrotyrosination of peptide residue p3Y induces a profound conformational change of the H-2Kb residue E152, altering TCR recognition The conformations of the peptides gp34 and NY-gp34 are very similar when bound to H-2Kb, besides a lateral shift of 0.7 A of the b side chain of the nitrotyrosine PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/22187127 p3NY in H-2K /NY-gp34 when compared to the conformation of p3Y in 
H-2Kb/gp34. In H-2Kb/gp34 the side-chain of p3Y projects down into the Dpocket formed by residues Q114, E152, R155, L156 and Y159. Three hydrogen bond interactions are formed between the side chain of p3Y and the side chains of the H-2Kb residues E152 and R155. Nitrotyrosination of p3Y alters only the conformation of the H-2Kb heavy chain residue E152 which, in order to accommodate the side-chain of p3NY, undergoes a large conformational modification in H-2Kb/ NY-gp34 when compared to H-2Kb/gp34. In contrast, the rest of the H-2Kb peptide-binding groove is not affected by the nitrotyrosination of the peptide in H-2Kb/NYgp34 when compared to H-2Kb/gp34. In conclusion, substitution of the tyrosine residue at position 3 of the peptide gp34 to a nitrotyrosine exclusively affects the conformation of the side-chain of the H-2Kb residue E152. The specificity of the T cell hybridoma 24H1 to H-2Kb/NY-gp34 thus seems to be dependent on the conformation of the MHC region surrounding the side-chains of p3NY and the H-2Kb residue E152. It should be noted that previous studies have demonstrated the importance of this region for recognition by H-2Kb-specific TCRs. One may thus speculate that nitrotyrosination of the immunodominant peptide which alters considerably T cell recognition may be advantageous for the virus in order to escape immune recognition. Nitrotyrosination of gp34 strains the overall conformation of the H-2Kb complex and significantly decreases its stability Two additional interactions are formed between peptide residue p3NY and the heavy chain residues Q114 and Y116 in the H2Kb/NY-gp34 peptide when compared to H-2Kb/gp34, suggesting that nitrotyrosination of the peptide may result in higher stabilization cap
