This study describes the use of the CD8/major histocompatibility complex (MHC) class I crystal structure as a template for the de novo design of low-molecular-weight surface mimetics. The analogs were designed from a local surface region on the CD8 a-chain directly adjacent to the bound MHC class I, to block the protein associations in the T-cell activation cluster that occur upon stimulation of the cytotoxic T lymphocytes (CTLs). One small conformationally restrained peptide showed dose-dependent inhibition of a primary allogeneic CTL assay while having no effect on the CD4-dependent mixed lymphocyte reaction (MLR). The analog's activity could be modulated through subtle changes in its side chain composition. Administration of the analog prevented CD8-dependent clearance of a murine retrovirus in BALB/c mice. In C57BL/6 mice challenged with the same retrovirus, the analog selectively inhibited the antiviral CTL responses without affecting the ability of the CTLs to generate robust allogeneic responses.