Determination of amino acids on agretopes of pigeon cytochrome c‐related peptides specifically bound to I‐A allelic products

Abstract

In our prior study it was demonstrated that residues 46 and 54 on a synthetic peptide, AEGFSYTVANKNKGIT (50V), work as an agretope (site contacts with major histocompatibility complex molecules) and residues 50 and 52 function as an epitope (site contacts with T cell receptor), when tri-molecular complexes are formed among 50V,I-Ab and the T cell receptor. 50V was composed of residues 43 to 58 of pigeon cytochrome c (p43-58) except that the aspartic acid (D) at residue 50 was substituted by valine (V). Substitution of agretopic residues on 50V changed this I-Ab-binding peptide to an I-Ak-binding peptide, suggesting that positions 46 and 54 work as an agretope in I-Ak-restricted T cell responses. In the present study we examined whether residues 46 and 54 of 50V worked as agretopes in T cell responses restricted to other I-A haplotypes. The 50V-related peptides with phenylalanine (F) at position 46 and alanine (A) at position 54 bound tightly to I-Ab, I-Ad, I-Aq and I-As molecules and stimulated T cells most potently in mice bearing these I-A haplotypes. In contrast, 50V-related peptides carrying D at position 46 and A at position 54 bound most potently to I-Ak molecules, and the peptides with arginine (R) at position 46 and A at position 54 bound most efficiently to I-Av molecules. The present findings, thus, demonstrate that the agretopic positions on the p43-58 related peptides are preserved in T cell responses restricted to each I-A haplotype studied, and that the specific amino acids on the agretopic positions exist a priori for each I-A allele-specific structure.

αβTCRs Differ in the Degree of Their Specificity for the Positively Selecting MHC/Peptide Ligand

“…We have produced mice expressing covalently bound analogues of Ep (Ep58K and Ep63K) and pigeon cytochrome c-derived peptide PCC (43)(44)(45)(46)(47)(48)(49)(50)(51)(52)(53)(54)(55)(56)(57)(58) , and TCR␣-chain knockout mice were purchased from The Jackson Laboratory (Bar Harbor, ME). All mice were housed under specific pathogen-free conditions in the animal care facility at the Medical College of Georgia (Augusta, GA).…”

“…All three hybridomas were also tolerant to covalent A b Ep complex and A b bound with self peptides. The first hybridoma, 2.1.4, recognized the PCC (43-58) peptide or its analogue PCC(50V54A), which had enhanced binding and stimulatory properties over the original PCC peptide (43). The TCR cloned from this CD4 ϩ T cell hybridoma was expressed in vivo using either the CD2 promoter (VA cassette) or genuine TCR promoters (pT␣/pT␤ cassettes) (32,33).…”

Interaction of pigeon cytochromec-(43–58) peptide analogs with either T cell antigen receptor or I-Ab molecule

“…1). Because it had been shown that P1 and P9 are critical sites for interaction with I-A b , analogs with substitutions of charged amino acids at P1 or P9 were not examined in the present experiments (7,8,12,13). Fig.…”

“…However, this orientation appears to differ from the model proposed by Jorgensen et al (6), in which the TCR is rotated approximately 90°relative to the peptide-MHC surface. Thus far, no crystal structure of trimolecular complex, including class II molecules, has been determined, and it is unclear whether formation of the trimolecular complex is predicted by a general model demonstrated with a MHC class I molecule or is determined individually by each of the components.In prior studies (7,8), it has been demonstrated that P1 of a pigeon cytochrome c-(43-58) peptide (p43-58) is a main anchor to the I-A molecule and a particular amino acid at the P1 determines allele-specific peptide binding. Hammer et al (9) and Jardetzky et al (10) also showed that the P1 residue of HLA-DR1-binding peptides represents the most important anchor.…”

“…In prior studies (7,8), it has been demonstrated that P1 of a pigeon cytochrome c-(43-58) peptide (p43-58) is a main anchor to the I-A molecule and a particular amino acid at the P1 determines allele-specific peptide binding. Hammer et al (9) and Jardetzky et al (10) also showed that the P1 residue of HLA-DR1-binding peptides represents the most important anchor.…”