Cytotoxic T (Tc)‐cell responses against influenza virus infection in BALB/c (H‐2^d) mice are dominated by Tc clones reactive to the viral nucleoprotein (NP). Here, we report investigations using recombinant vaccinia viruses (VV) encoding major histocompatibility complex (MHC) class I H‐2K^d molecules differing by a single amino acid from glutamine (wild‐type, K^dw) to histidine (mutant, K^dm) at position 114 located in the floor of the peptide‐binding groove. Influenza‐infected target cells expressing K^dw were strongly lysed by K^d‐restricted Tc cells against A/WSN influenza virus or the immunodominant peptide of viral NP (NPP^147–155), whereas infected K^dm‐expressing targets gave little or no lysis, respectively, thus showing the immunodominance of NPP^147–155. K^dm‐expressing target cells saturated with synthetic NPP^147–155 (10⁻⁵ m) were lysed similarly to K^dw‐expressing targets by NPP^147–155‐specific Tc cells. Thus the defect in influenza‐infected K^dm‐expressing targets was quantitative; insufficient K^dm–peptide complexes were expressed. Tc‐cell responses against four other viruses or alloantigens showed no effect of K^dm. When peptide transport‐defective cells were infected with VV‐K^dw or VV‐K^dm and co‐infected with a recombinant VV encoding an endoplasmic reticulum‐targeted viral peptide, two influenza haemaglutinin peptides caused higher expression of K^dw than NPP^147–155 indicating their higher affinity for K^dw. These results are inconsistent with the hypothesis that immunodominance in the anti‐influenza response reflects high affinity of the immunodominant peptide, but are consistent with skewing of the Tc‐cell receptor repertoire.