Lipopeptide ligands captured by MHC class I molecules undergo dynamic conformational changes that affect their...

A fraction of the major histocompatibility complex class I proteins can bind N-myristoylated short lipopeptides rather than conventional long peptides. The molecular mechanisms underlying lipopeptide antigen presentation were recently delineated for N-myristoylated 4-mer lipopeptides (C14-Gly1-Gly2-Ala3-Ile4; C14nef4) derived from the retroviral Nef protein. The C14nef4 lipopeptides are captured by the rhesus major histocompatibility complex class I allomorph, Mamu-B∗05104, and recognized by specific αβ T-cell receptors (TCRs). The crystal structure of the Mamu-B∗05104-C14nef4-TCR complex indicates that both ends of C14nef4, namely, myristic acid and C-terminal Ile4, are anchored at the antigen-binding groove, leaving Gly1, Gly2, and Ala3 exposed. Among these residues, only the amide bond of Gly1 forms a hydrogen bond with TCRs and serves as a primary T-cell epitope. However, it remains unclear how antigenic and nonantigenic lipopeptides exist, both of which share the primary T-cell epitope. To gain insight into this enigma, we utilized C14nef4 and its analogs with an amino acid substitution for Ala3. Biolayer interferometry experiments with immobilized TCRs and lipopeptide-bound Mamu-B∗05104 indicated that the antigenic strength varied among these lipopeptides. The crystal structures of Mamu-B∗05104 complexed with either C14nef4 or each of its five analogs showed a downward shift in the proximal part (C₁-C₄ carbons) of the hydrocarbon chain and the linked Gly1 residue for poorly antigenic analogs. Furthermore, molecular dynamics simulations indicated that lipopeptide ligands alter their conformation dynamically, with differential efficiency in exposing Gly1 externally. Thus, the antigenic strength of lipopeptides is affected by their intrinsic ability to sustain a T-cell epitope-exposed configuration.