Earlier mutational studies [40
] have demonstrated a crucial role for several CD1d residues for i
NKT cell activation. All of these residues have now been shown through structural analysis to either participate in the hydrogen-bond network that stabilizes the antigenic head group of the glycolipid for proper TCR engagement (Arg79, Asp80, Asp153 and Thr156, see ), and/or they are predicted to directly interact with the TCR (Glu83, Arg79) to facilitate CD1d-lipid antigen-TCR ternary complex formation [40
]. Asp80 in the α1-helix interacts with the 3-OH group of the sphingosine base, whereas Asn153 in mouse CD1d (151 in human CD1d) stabilizes the head groups through interaction with either the 2′- and/or the 3′-OH groups of the sugars. Thr156 mouse CD1d (Thr154 in human CD1D) interacts with the oxygen of the O
-glycosidic linkage and with the sphingosine backbone nitrogen. Additional residues, such Arg79 in mouse CD1d, can provide additional specificity for the ligand, but are also in a suitable location for interacting with the incoming TCR
Fig. 4 CD1d hydrogen-bonding network. The CD1d ligands (yellow, green and cyan) are stabilized by hydrogen-bonds (dashed lines) between the α1 and α2-helices of either human CD1d (A) or mouse CD1d (B). (A) Comparison of α-GalCer presentation (more ...)
Interestingly, mouse and human CD1d are very similar in structure and in their binding of α-GalCer, and in fact there is interspecies cross reactivity between mouse and human i
NKT cells [41
]. The only major difference occurs around Trp153 in the human ortholog, which raises and tilts the galactose head group slightly, whereas the corresponding mouse residue is the much smaller glycine, which does not influence ligand binding ().
Both α-GalCer and the Sphingomonas glycolipid GalA-Gsl activate iNKT cells, but α-GalCer is a much more potent antigen. Most of the polar interactions between the two ligands and mouse CD1d residues are conserved; however, slight structural differences, such as the lack of the sphingosine 4-OH group in GalA-Gsl, affect the fine positioning of the ligand in the binding groove. Compared to α-GalCer, GalA-Gsl lacks the hydrogen bond with Asp80 and, as a result, the sphingosine chain is inserted slightly deeper into the F′ pocket (). This altered interaction results in an overall tilt of GalA-Gsl in the binding groove, which leads to lateral shift of the galacturonosyl head group by about 1 Å along the CD1d surface. This tilt in turn could lead to a different interaction with the TCR and, hence, could explain the weaker T cell stimulation. α-GalCer also induces slight, but potentially important structural changes in the α1-helix, which cause a more intimate association with the ligand (). It was proposed that the two hydrogen bonds between the 3-OH and 4-OH of the sphingosine base of the ceramide lipid of α-GalCer and CD1d amino acid Asp80 are responsible for pulling the α1-helix toward the ligand. The induced structural changes result in the formation of a roof above the F′ pocket, which increases the T cell recognition surface and, could provide additional avidity for the TCR.