The Structure and Stability of an HLA-A*0201/Octameric Tax Peptide Complex with an Empty Conserved Peptide-N-Terminal Binding Site

Abstract

The crystal structure of the human class I MHC molecule HLA-A2 complexed with of an octameric peptide, Tax8 (LFGYPVYV), from human T cell lymphotrophic virus-1 (HTLV-1) has been determined. This structure is compared with a newly refined, higher resolution (1.8 Å) structure of HLA-A2 complexed with the nonameric Tax9 peptide (LLFGYPVYV) with one more N-terminal residue. Despite the absence of a peptide residue (P1) bound in the conserved N-terminal peptide-binding pocket of the Tax8/HLA-A2 complex, the structures of the two complexes are essentially identical. Water molecules in the Tax8 complex replace the terminal amino group of the Tax9 peptide and mediate a network of hydrogen bonds among the secondary structural elements at that end of the peptide-binding groove. Thermal denaturation measurements indicate that the Tax8 complex is much less stable, ΔTm = 16°C, than the Tax9 complex, but both can sensitize target cells for lysis by some Tax-specific CTL from HTLV-1 infected individuals. The absence of a P1 peptide residue is thus not enough to prevent formation of a “closed conformation” of the peptide-binding site. TCR affinity measurements and cytotoxic T cell assays indicate that the Tax8/HLA-A2 complex does not functionally cross-react with the A6-TCR-bearing T cell clone specific for Tax9/HLA-A2 complexes.

Selective activation, expansion, and monitoring of human iNKT cells with a monoclonal antibody specific for the TCR α‐chain CDR3 loop

“…In order to raise antibodies against the human invariant Va24-Ja18 TCR, a cyclic peptide representing the predicted CDR3 loop of the invariant TCRa sequence [CH2CO-CVVSDRGSTLGRLAD-C (thioether linkage to CH 2 )-G-COOH] was modeled using previous TCR crystal structures as templates [48,49]. The peptide was cyclized in situ using a thioether linkage, followed by deprotection of the N-terminal Cys residue and release from the resin for purification (SynPep, Dublin, CA).…”

Structures of MART-126/27–35 Peptide/HLA-A2 Complexes Reveal a Remarkable Disconnect between Antigen Structural Homology and T Cell Recognition

“…12,21 Furthermore, the prediction that the AAG nonamer would adopt the bulged conformation is inconsistent with the database of known peptide/HLA-A2 structures, as this would require the peptide to bind with an empty P1 pocket, a binding mode that has not been observed with nonameric peptides, even when the peptide has a sub-optimal P2 anchor. 27 Here, we report the structure of the native MART-1 [27][28][29][30][31][32][33][34][35] AAG nonamer and a number of variants bound to HLA-A2. Inconsistent with earlier predictions, the native peptide does not adopt the bulged conformation of the ELA decamer.…”

“…7,27 Crystals of the A1L-modified MART-1 [27][28][29][30][31][32][33][34][35] nonamer (LAG nonamer), the A2L-modified nonamer (ALG nonamer), and the native MART-1 [26][27][28][29][30][31][32][33][34][35] decamer (EAA decamer) complexed with HLA-A2 grew readily from these conditions, although subtle modifications resulted in improved crystals. All peptides studied are shown in Table 1.…”

“…Structures of MART-1 26/27-35 -based peptide/HLA-A2 complexes reveal bulged and extended peptide conformations Crystals of the native MART-1 [27][28][29][30][31][32][33][34][35] nonamer (AAG nonamer) complexed with HLA-A2 were grown by screening conditions previously used to crystallize peptide/HLA-A2 complexes in our laboratory. 7,27 Crystals of the A1L-modified MART-1 [27][28][29][30][31][32][33][34][35] nonamer (LAG nonamer), the A2L-modified nonamer (ALG nonamer), and the native MART-1 [26][27][28][29][30][31][32][33][34][35] decamer (EAA decamer) complexed with HLA-A2 grew readily from these conditions, although subtle modifications resulted in improved crystals.…”

“…Rather, the peptide adopts an extended conformation nearly identical with that of the ALG nonamer. To further investigate, we determined the structures of the native MART-1 [26][27][28][29][30][31][32][33][34][35] EAA decamer and the 27-35 Ala2→Leu ALG nonamer bound to HLA-A2 at a higher resolution to closely examine the dynamics of the peptide previously described by Sliz et al 24 We also determined the structure of another variant, the 27(L)-35 nonamer (LAGIGILTV; LAG nonamer) that has been proposed as an improved melanoma vaccine candidate. 28 All MART-1 peptides and variants studied are shown in Table 1.…”