ate the side chain of p3NY, thus resulting in unfavorable strained conformations. Besides straining the backbone of the heavy chain residues 124126 and 156157, the side chain of p3NY also incurred strained higher internal energy conformations for the side-chains of residues V97, I98 and Q114, when compared to H-2Kb/gp34. In conclusion, the potential gain in enthalpy resulting from the formation of two additional interactions between p3NY and the H-2Kb residues Q114 and Y116 may be outweighed by the loss of a hydrogen bond interaction formed with the side chain of E152 combined with the free energy cost resulting from the strained conformations of several MHC residues. Our results indicate that formation of additional interactions between peptides and MHC residues does not always necessarily increase the overall stabilization of MHC complexes. Furthermore, alteration of MHC complex stabilization through nitrotyrosination of the presented epitope may represent an additional alternative strategy used by the virus to prevent adequate presentation of the peptide at the surface of the infected cell. Nitrotyrosination of p4Y in H-2Db/NY-gp33 directly affects recognition by H-2Db/gp33-specific TCRs The previously determined crystal structures of H-2Db and H2Kb in complex with gp33 demonstrated that the peptide binds in two diametrically opposed manners to the two MHC class I molecules. Thus in contrast to H-2Kb/gp33 and H-2Kb/ gp34, in which the side-chain of the tyrosine residue acts as a secondary anchor position, the side-chain of p4Y in H-2Db/gp33 protrudes out of the peptide-binding cleft, playing a key role in TCR recognition. Indeed, mutation of p4Y to a MHC-I-Restricted Nitrotyrosinated Neoantigen 7 MHC-I-Restricted Nitrotyrosinated Neoantigen Concluding remarks Our study provides a structural platform underlying the effects of peptide nitrotyrosination on initiation of TCR responses. Such PTMs PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/22188400 can directly affect TCR recognition by modifying the properties of key TCR-interacting residues on the presented peptide or by altering the conformations of other MHC residues that are of importance for TCR recognition. Nitrotyrosination can also indirectly affect TCR recognition by severely destabilizing the MHC complex. Although additional interactions were formed between the PTM peptide and H-2Kb, the conformation of a large amount of MedChemExpress 763113-22-0 surrounding residues was strained, significantly reducing the overall stability of the MHC complex. We have previously demonstrated that subtle modifications in MHCrestricted peptides can result in significant alterations of MHC stabilization, a phenomenon that should be taken into account and measured upon trying to design altered peptide ligands for modulation of TCR response. Several other amino acid modifications caused by oxidative stress could result in similar effects on TCR recognition. In conclusion, our structural study indicates that the impact of post-translational modifications could be dual, possibly allowing for viral immune escape from TCR recognition but also potentially inducing the 
expansion of subset of T cells that could induce autoreactivity. Likewise, nitrotyrosination of self-peptides during periods of inflammation or oxidative stress could lead to the formation of neo-epitopes that, through the same mechanisms described herein for gp33/34, could escape the constraints of central tolerance. The impact of these modified de-novo MHC complexes on the initiation of unwanted T cell responses th