Changes in the repertoire of peptides bound to HLA-B27 subtypes and to site-specific mutants inside and outside pocket B.

Abstract

SummaryHLA-B27 subtypes share many structural features, including their pocket ]3, which interacts with a conserved Arg residue at the second position of B*2705-bound peptides. Subtypes differ among each other at other locations in the peptide binding site. In this study, metabolic labeling and radiochemical pool sequencing were used to address the following issues: (a) presence of the Arg 2 (R2) motif among peptides bound to the various HLA-B27 subtypes; (b) influence of mutations inside and outside pocket B on this motif; and (c) the degree of similarity among the peptide pools bound to the various B27 subtypes. Sequencing of Arg-labeled peptide pools extracted from B'2701 to B'2706, and from two site-directed mutants ofB'2705 with changes outside pocket B, indicated that all of these molecules bind peptides with Arg at position 2. Peptides from several mutants with changes altering the structure of pocket 13, and from one mutant at the pocket B rim, also retained the lk2 motif. However, this was absent in the peptide pool extracted from the M45 mutant, in which the negative charge of pocket B, conferred to HLA-B27 by Glu45, was canceled. These results indicate that alterations outside pocket B, and even disruption of the network of hydrogen bonds that stabilizes Arg binding in pocket B, do not impair binding of peptides bearing the R2 motif, but a nonconservative substitution at position 45 does. As a substantial fraction of anti-B ~ cytotoxic T lymphocyte (CTL) clones crossreact with the M45 mutant (Villadangos, J., B. Galocha, D. L6pez, V. Cairo, andJ.A. L6pez de Castro. 1992.J. Irnraunol. 149:505) this result suggests that determinant mimicry by nonidentical peptides may frequently account for unexpected CTL crossreactions. Metabolic labeling with various other amino acids and radiochemical sequencing revealed similarities, but also substantial differences, among the peptide pools from the various HLA-B27 subtypes. This strongly suggests that many peptides bind to multiple subtypes, but significant subsets of peptides bound to a given HLA-B27 subtype do not bind to other subtypes or do so with greatly altered effidency. These results indicate the importance of polymorphism outside pocket B in modulating peptide binding to HLA-B27.

Differences in endogenous peptides presented by HLA-B*2705 and B*2703 allelic variants. Implications for susceptibility to spondylarthropathies.

“…C1R-B*2705 (B*27052), B*2702, and B*2703 were kindly provided by Dr. J.A. Lopez de Castro (17). For peptide elution, they were cultured in roller bottles in RPMI 1640 medium/10% FCS/ 2 mM glutamine/ penicillin G at 50 U/ml and streptomycin at 50 g/ml.…”

Enhanced intracellular replication of Salmonella enteritidis in HLA–B27–expressing human monocytic cells: Dependency on glutamic acid at position 45 in the B pocket of HLA–B27

“…HC homodimerization also appears to occur distal to the ER and/or during HC recycling from the cell surface (11,12) in a process that is associated with loss of ␤ 2 -microglobulin and/or peptide, and it appears to be distinct from ER misfolding. Altering amino acids that comprise the B pocket, a region in the peptide-binding groove that also has a dominant influence on peptide selection (13)(14)(15), can correct misfolding. For example, substitution of methionine for glutamic acid at position 45, either alone or in combination with other changes, dramatically enhances HLA-B27 HC folding and prevents ER dimerization (10,11).…”

The impact of HLA‐B micropolymorphism outside primary peptide anchor pockets on the CTL response to CMV

“…In many cases, such minor sequence differences occur within the primary peptide anchor pockets and, consequently, major differences in MHCpeptide binding specificity provide the obvious evolutionary pressure that maintains these allelic dimorphisms within populations [35][36][37]. Natural class I polymorphism outside the B and F anchor pockets modulates peptide ligand specificity much more subtly, as shown by studies comparing self-peptide presentation [24,[38][39][40][41].…”