This study describes an analysis of the interaction of individual
amino acid residues of the
vesicular stomatitis virus (VSV) nucleocapsid antigenic octapeptide (N52-59;
Arg-Gly-Tyr-Val-Tyr-Gln-Gly-Leu) with the H-2Kb molecule and
T-cell receptors (TCRs). Tyr-3, Tyr-5, and Leu-8 were the positions in the
peptide found to be H-2Kb contact residues by analyzing single
alanine-substituted
peptides in a competition assay with a Kb-restricted antigenic nonapeptide of Sendai virus. Arg-1, Gly-2, Val-4, Gln-6, and Gly-7 of the
peptide were identified as putative TCR contact residues by testing the
peptide analogs for their capacity to sensitize targets for VSV-specific cytolytic T-lymphocyte clones. The octamer N52-59 was the optimal length of the
peptide required for binding to Kb. This
peptide length requirement and the finding of an irregular interspersing of major histocompatibility complex and TCR contact residues are most consistent with the conclusion that the
peptide is in an extended conformation in the
antigen binding groove. Furthermore, data on binding of truncated
peptides show that, although the Arg-1 side chain has been assigned as a TCR contact residue, the main-chain atoms of the N-terminal amino group are most likely involved in interacting with the major histocompatibility complex molecule. A panel of H-2Kb point mutants was constructed to explore the effect of altered
amino acid residues on the binding of N52-59. Mutants with amino acid substitutions along the floor of the groove all bound the VSV
peptide but modulated its interaction with Kb, apparently causing subtle changes in the spatial arrangement of some specific TCR contact residues in the
peptide.