Peptide Recognition by Two HLA-A2/Tax 11–19 -Specific T Cell Clones in Relationship to Their MHC/Peptide/TCR Crystal Structures 1 Stefan Hausmann,* William E. Biddison, ² Kathrine J. Smith, ‡§ Yuan-Hua Ding, § David N. Garboczi, ¶ Ursula Utz,  Don C. Wiley, ‡§ and Kai W. Wucherpfennig 2 * The crystal structures of two human TCRs specific for a HTLV-I Tax peptide bound to HLA-A2 were recently determined, for the first time allowing a functional comparison of TCRs for which the MHC/peptide/TCR structures are known. Extensive amino acid substitutions show that the native Tax residues are optimal at each peptide position. A prominent feature of the TCR contact surface is a deep pocket that accommodates a tyrosine at position 5 of the peptide. For one of these TCRs, this pocket is highly specific for aromatic residues. In the other TCR structure, this pocket is larger, allowing many different residues to be accom- modated. The CTL clones also show major differences in the specificity for several other peptide residues, including side chains that are not directly contacted by the TCR. Despite the specificity of these clones, peptides that are distinct at five or six positions from Tax 11–19 induce CTL activity, indicating that substantial changes of the peptide surface are tolerated. Human peptides with limited sequence homology to Tax 11–19 represent partial TCR agonists for these CTL clones. The distinct functional properties of these CTL clones highlight structural features that determine TCR specificity and cross-reactivity for MHC-bound peptides. The Journal of Immunology, 1999, 162: 5389 –5397. T he specificity of T cell-mediated immune responses is de- termined by TCR recognition of antigenic peptides bound to MHC class I or class II molecules. Studies performed over the past several years have demonstrated that the same TCR can recognize peptides that are quite distinct in their primary se- quence from each other. Such cross-reactivity of TCRs with dis- tinct MHC/peptide complexes is important for thymic selection and survival of mature T cells, and has been implicated in trans- plant rejection and autoimmune diseases (1–9). The MHC/peptide/ TCR crystal structures that have been determined allow structural features to be analyzed that contribute to the specificity and de- generacy of TCR recognition. High resolution MHC/peptide/TCR structures have been deter- mined for two human TCRs (A6 and B7) that are specific for an immunodominant HTLV-I Tax peptide bound to HLA-A2 (10, 11). A similar diagonal binding mode of the TCR has been observed for these two human complexes and a murine MHC/peptide/TCR com- plex (10 –12). The diagonal binding mode buries most of the peptide in the MHC class I/TCR complex and allows the flat TCR surface to interact with the peptide by fitting down between the highest points of the MHC helixes. These crystal structures as well as mutagenesis experiments with murine MHC class I- and class II-restricted TCRs indicate that the diagonal binding mode may be general (13, 14). The structures of A6 and B7 TCRs demonstrate how different TCRs recognize the same MHC/peptide complex (10, 11). The surface chemistries of the two TCRs are different, since only one of the 17 contact residues of the B7 TCR with the HLA-A2/Tax complex is identical with the A6 TCR. Nevertheless, the same general binding mode is observed, with a certain degree of rotation and tilt of the V domains. The V domain of B7 is rotated about 10° counterclockwise relative to the A6 TCR and is tipped closer to the MHC molecule. Smaller differences are observed in the position of the V domains. In the A6 structure, the CDR1 and CDR3 loops of both V and V contact the Tax 11–19 peptide. The CDR1 loop of  is posi- tioned over the peptide N-terminus, while the CDR1 loop of  is positioned over the C-terminal end of the peptide. A tyrosine at position 5 of the Tax peptide is bound in a deep pocket at the center of the TCR that is shaped by the CDR3 loops of both  and . The CDR2 loops of V and V are positioned over the helixes of the 2 and 1 domains of the MHC molecule, respectively. In the A6 and B7 structures, substantial TCR contacts are only made to residues Y5 and Y8 of the Tax peptide. Also, the total TCR contact surface with the peptide is relatively small, approximately one-third of the total TCR contact surface with the MHC/peptide complex (326 Å 2 of 998 Å 2 ). These observations raise the following questions. 1) Which structural features contribute to the specificity of these TCRs for the HLA-A2/Tax peptide complex? 2) How different are these CTL clones in their fine specificity? 3) To what extent can the peptide surface that is contacted by these TCRs be modified? *Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Bos- ton, MA 02115; ² Molecular Immunology Section, Neuroimmunology Branch, Na- tional Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892; ‡ Laboratory of Molecular Medicine, Department of Medicine, The Children’s Hospital, Howard Hughes Medical Institute, Boston, MA 02115; § De- partment of Molecular and Cellular Biology, Harvard University, Howard Hughes Medical Institute, Cambridge, MA 02138; ¶ Structural Biology Section, National In- stitute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, MD 20852; and  Instiut de Recherches Cliniques de Montre ´al, Laboratoire d’Immunologie, Montreal, Quebec, Canada Received for publication November 3, 1998. Accepted for publication February 8, 1999. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 This work was supported by the National Institutes of Health (Grant AI42316 to K.W.W. and Grant HD-17461 to D.C.W.), the Howard Hughes Medical Institute, a fellowship from the Deutscher Akademischer Austauschdienst (to S.H.), and a Well- come Trust Traveling Fellowship (to K.J.S.). K.W.W. is a Harry Weaver Neuro- science Scholar of the National Multiple Sclerosis Society. D.C.W. is an investigator with the Howard Hughes Medical Institute. 2 Address correspondence and reprint requests to Dr. Kai W. Wucherpfennig, De- partment of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, 44 Binney St., Boston, MA 02115. E-mail address: wucherpf@mbcrr.harvard.edu Copyright © 1999 by The American Association of Immunologists 0022-1767/99/$02.00