Design of natural killer T cell activators: structure and function of a microbial glycosphingolipid bound to mouse CD1d.
Natural killer T (NKT) cells provide an innate-type immune response upon T cell receptor interaction with CD1d-presented antigens. We demonstrate through equilibrium tetramer binding and antigen presentation assays with Valpha14i-positive NKT cell hybridomas that the Sphingomonas glycolipid alpha-galacturonosyl ceramide (GalA-GSL) is a NKT cell agonist that is significantly weaker than alpha-galactosylceramide (alpha-GalCer), the most potent known NKT agonist. For GalA-GSL, a shorter fatty acyl chain, an absence of the 4-OH on the sphingosine tail and a 6'-COOH group on the galactose moiety account for its observed antigenic potency. We further determined the crystal structure of mCD1d in complex with GalA-GSL at 1.8-A resolution. The overall binding mode of GalA-GSL to mCD1d is similar to that of the short-chain alpha-GalCer ligand PBS-25, but its sphinganine chain is more deeply inserted into the F' pocket due to alternate hydrogen-bonding interactions between the sphinganine 3-OH with Asp-80. Subsequently, a slight lateral shift (>1 A) of the galacturonosyl head group is noted at the CD1 surface compared with the galactose of alpha-GalCer. Because the relatively short C(14) fatty acid of GalA-GSL does not fully occupy the A' pocket, a spacer lipid is found that stabilizes this pocket. The lipid spacer was identified by GC/MS as a mixture of saturated and monounsaturated palmitic acid (C(16)). Comparison of available crystal structures of alpha-anomeric glycosphingolipids now sheds light on the structural basis of their differential antigenic potency and has led to the design and synthesis of NKT cell agonists with enhanced cell-based stimulatory activities compared with alpha-GalCer.
Department of Chemistry and Molecular Biology and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Douglass
D
Wu
Dirk M
DM
Zajonc
Masakazu
M
Fujio
Barbara A
BA
Sullivan
Yuki
Y
Kinjo
Mitchell
M
Kronenberg
Ian A
IA
Wilson
Chi-Huey
CH
Wong
D000818:Animals; D018949:Antigens, CD1 / Q000737:chemistry / Q000378:metabolism*; D055610:Antigens, CD1d; D001665:Binding Sites; D006028:Glycosphingolipids / Q000737:chemistry* / Q000378:metabolism*; D006860:Hydrogen Bonding; D066298:In Vitro Techniques; D007694:Killer Cells, Natural / Q000276:immunology* / Q000378:metabolism*; D008213:Lymphocyte Activation; D046911:Macromolecular Substances; D051379:Mice; D008958:Models, Molecular; D015394:Molecular Structure; D020836:Protein Structure, Quaternary; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism*
D018949:Antigens, CD1; D055610:Antigens, CD1d; D006028:Glycosphingolipids; D046911:Macromolecular Substances