The CD1 family is a large cluster of non-polymorphic, major histocompatibility complex (MHC) class-I-like molecules that bind distinct lipid-based antigens that are recognized by T cells. The most studied group of T cells that interact with lipid antigens are natural killer T (NKT) cells, which characteristically express a semi-invariant T-cell receptor (NKT TCR) that specifically recognizes the CD1 family member, CD1d. NKT-cell-mediated recognition of the CD1d-antigen complex has been implicated in microbial immunity, tumour immunity, autoimmunity and allergy. Here we describe the structure of a human NKT TCR in complex with CD1d bound to the potent NKT-cell agonist alpha-galactosylceramide, the archetypal CD1d-restricted glycolipid. In contrast to T-cell receptor-peptide-antigen-MHC complexes, the NKT TCR docked parallel to, and at the extreme end of the CD1d-binding cleft, which enables a lock-and-key type interaction with the lipid antigen. The structure provides a basis for the interaction between the highly conserved NKT TCR alpha-chain and the CD1d-antigen complex that is typified in innate immunity, and also indicates how variability of the NKT TCR beta-chain can impact on recognition of other CD1d-antigen complexes. These findings provide direct insight into how a T-cell receptor recognizes a lipid-antigen-presenting molecule of the immune system.

Natural Killer T (NKT) cells respond to a variety of CD1d-restricted antigens (Ags), although the basis for Ag discrimination by the NKT cell receptor (TCR) is unclear. Here we describe NKT TCR fine specificity against several closely related Ags, termed altered glycolipid ligands (AGLs), which differentially stimulate NKT cells. The structures of five ternary complexes all revealed similar docking. Acyl chain modifications did not affect the interaction, but reduced NKT cell proliferation, indicating an affect on Ag processing or presentation. Conversely, truncation of the phytosphingosine chain caused an induced fit mode of TCR binding that affected TCR affinity. Modifications in the glycosyl head group had a direct impact on the TCR interaction and associated cellular response, with ligand potency reflecting the t1/2 life of the interaction. Accordingly, we have provided a molecular basis for understanding how modifications in AGLs can result in striking alterations in the cellular response of NKT cells.

Although it has been established how CD1 binds a variety of lipid antigens (Ag), data are only now emerging that show how αβ T cell receptors (TCRs) interact with CD1-Ag. Using the structure of the human semiinvariant NKT TCR–CD1d–α-galactosylceramide (α-GalCer) complex as a guide, we undertook an alanine scanning mutagenesis approach to define the energetic basis of this interaction between the NKT TCR and CD1d. Moreover, we explored how analogues of α-GalCer affected this interaction. The data revealed that an identical energetic footprint underpinned the human and mouse NKT TCR–CD1d–α-GalCer cross-reactivity. Some, but not all, of the contact residues within the Jα18-encoded invariant CDR3α loop and Vβ11-encoded CDR2β loop were critical for recognizing CD1d. The residues within the Vα24-encoded CDR1α and CDR3α loops that contacted the glycolipid Ag played a smaller energetic role compared with the NKT TCR residues that contacted CD1d. Collectively, our data reveal that the region distant to the protruding Ag and directly above the F′ pocket of CD1d was the principal factor in the interaction with the NKT TCR. Accordingly, although the structural footprint at the NKT TCR–CD1d–α-GalCer is small, the energetic footprint is smaller still, and reveals the minimal requirements for CD1d restriction.