To elucidate the structural basis of T cell recognition of hapten- modified antigenic peptides, we studied the interaction of the T1 T cell antigen receptor (TCR) with its ligand, the H-2K(d)-bound Plasmodium berghei circumsporozoite peptide 252-260 (SYIPSAEKI) containing photoreactive 4- azidobenzoic acid (ABA) on P. berghei circumsporozoite Lys259. The photoaffinity-labeled TCR residue(s) were mapped as Tyr48 and/or Tyr50 of complementary determining region 2β (CDR2β). Other TCR-ligand contacts were identified by mutational analysis. Molecular modeling, based on crystallographic coordinates of closely related TCR and major histocompatibility complex I molecules, indicated that ABA binds strongly and specifically in a cavity between CDR3α and CDR2β. We conclude that TCR expressing selective Vβ and CDR3α sequences form a binding domain between CDR3α and CDR2β that can accommodate nonpeptidic moieties conjugated at the C-terminal portion of peptides binding to major histocompatibility complex (MHC) encoded proteins.