Structure and Peptidomes of Swine MHC Class I with Long Peptides Reveal the Cross-Species Characteristics of the Novel N-Terminal Extension Presentation Mode

Abstract

Antigenic peptide presentation by the MHC is essential for activating T cells. The current view is that the peptide termini are tethered within the closed Ag-binding groove of MHC class I (MHC-I). Recently, the N-terminal extension mode of peptide presentation has been observed in human MHC-I (HLA-I). In this study, we found that the N terminus of the long peptide can extend beyond the groove of swine MHC-I (SLA-1*0401), confirming that this phenomenon can occur across species. Removal of the N-terminal extra (P-1) residue of the RW12 peptide significantly reduced the folding efficiency of the complex, but truncation of the second half of the peptide did not. Consistent with previous reports, the second (P1) residue of the peptide is twisted, and its side chain is inserted into the A pocket to form two hydrogen bonds with polymorphic E63 and conserved Y159. Mutations of E63 disrupt the binding of the peptide, indicating that E63 is necessary for this peptide-binding mode. Compared with W167, which exists in most MHC-Is, SLA-I–specific S167 ensures an open N-terminal groove of SLA-1*0401, enabling the P-1 residue to extend from the groove. In this MHC class II–like peptide-binding mode, the A pocket is restrictive to the P1 residue and is affected by the polymorphic residues. The peptidomes and refolding data indicated that the open N-terminal groove of SLA-1*0401 allows one to three residues to extend out of the Ag-binding groove. These cross-species comparisons can help us better understand the characteristics of this N-terminal extension presentation mode.

Peptidomes and Structures Illustrate How SLA-I Micropolymorphism Influences the Preference of Binding Peptide Length

“…Structural analysis of SLA-1*13:01 and SLA-1*04:01 complexed with long peptides is helpful to clarify their different preferences for peptide length. We have previously identified a dodecapeptide (RW12) that can bind to SLA-1*04:01 and solved the structure of their complex pSLA-1*04:01 RW12 (13).…”

“…A random peptide repertoire was synthesized with a distribution of 19 amino acids other than cysteine at an equal molar ratio in every position, as previously reported ( 13 , 29 , 30 ). In brief, random peptide libraries of different lengths were synthesized separately according to this method and mixed in an equimolar ratio according to the average molecular mass before use.…”

“…The conformational differences of peptides with different lengths will further affect the recognition of the TCR repertoire (6). However, some recent studies found that some long peptides can extend from the N-or C-end of the peptide binding groove, which reveals a new pattern of MHC-I presenting long peptides (9)(10)(11)(12)(13). However, compared with short peptides, the number of identified long peptides is much lower, which limits the in-depth study of the mechanism of MHC-I presenting long peptides.…”

“…In recent years, the peptide length preference of MHC-I and its influence on the immune response have attracted increasing attention (5,14,15). The development of peptidomics provides powerful tools for accurately identifying the peptide binding characteristics of MHC-I, including peptide length preference (4,13,(16)(17)(18)(19). Mass spectrometry (MS) analysis of the human MHC-I (HLA-I) peptide ligand shows that some HLA-I alleles prefer to present longer peptides (5,14,15,20).…”

“…In addition, we also found a dodecapeptide RVEDVTNTAEYW (RW12 for short) which can bind to SLA-1*04:01. The complex structure (pSLA-1*04:01 RW12 ) indicated that the first Arg of RW12 can extend from the N terminus of the peptide binding groove of SLA-1*04:01 ( 13 ). In this study, the length distributions of peptides presented by SLA-I*04:01 and SLA-1*13:01 were compared by RPLD-MS, and it was found that SLA-1*13:01 can bind more long peptides than SLA-1*04:01.…”