Endogenous phosphatidylcholine and a long spacer ligand stabilize the lipid-binding groove of CD1b

CD1 proteins present lipid antigens to T cells. The antigens are acquired in the endosomal compartments. This raises the question of how the large hydrophobic CD1 pockets are preserved between the moment of biosynthesis in the endoplasmic reticulum and arrival to the endosomes. To address this issue, the natural ligands associated with a soluble form of human CD1b have been investigated. Using isoelectric focusing, native mass spectrometry and resolving the crystal structure at 1.8 A resolution, we found that human CD1b is simultaneously associated with endogenous phosphatidylcholine (PC) and a 41-44 carbon atoms-long spacer molecule. The two lipids appear to work in concert to stabilize the CD1b groove, their combined size slightly exceeding the maximal groove capacity. We propose that the spacer serves to prevent binding of ligands with long lipid tails, whereas short-chain lipids might still displace the PC, which is exposed at the groove entrance. The data presented herein explain how the CD1b groove is preserved, and provide a rationale for the in vivo antigen-binding properties of CD1b.