title: Characterization of peptides bound to extracellular and intracellular HLA-DR1 molecules.
abstract: Exogenous antigens are internalized by antigen-processing cells and processed within vesicular compartments to produce antigenic peptides that bind to newly synthesized MHC II molecules. These MHC class II peptide complexes are displayed at the plasma membrane and stimulate specific CD4+ T cells. In the present study, we established a method to isolate intracellular MHC molecules in a preparative scale (2-3 mg HLA-DR1) from endosomal compartments by Percoll density-gradient centrifugation. Peptides associated with HLA-DR1 in these intracellular fractions were released, purified by microbore HPLC, characterized by sequencing, and compared with the amino acid composition of peptides derived from MHC class II molecules obtained by solubilization of the plasma membrane. The binding affinity of these MHC fractions was analyzed by our highly sensitive binding assay using different DR1-restricted IM and Ii peptides. The results indicate that (a) intracellular MHC molecules show higher peptide-binding capacity, (b) peptides that are about 18-25 amino acids long need only a core region of 11 amino acids for binding, (c) specific positions of the peptides are important for DR1 binding, (d) most of the naturally processed peptides show a proline at position 2 or 3 that may represent a stop signal for trimming, and (e) Ii peptides are very abundant in DR1 peptide pools derived from intracellular compartments.
authors.affiliation: Medicine and Natural Sciences Research Center, University Tübingen, Germany.
authors.forename: H
authors.initials: H
authors.lastname: Max
authors.forename: T
authors.initials: T
authors.lastname: Halder
authors.forename: H
authors.initials: H
authors.lastname: Kropshofer
authors.forename: M
authors.initials: M
authors.lastname: Kalbus
authors.forename: C A
authors.initials: CA
authors.lastname: Müller
authors.forename: H
authors.initials: H
authors.lastname: Kalbacher
mesh_terms: D000595:Amino Acid Sequence; D002461:Cell Line, Transformed; D002499:Centrifugation, Density Gradient; D015798:HLA-DR1 Antigen / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000276:immunology; D013347:Subcellular Fractions / Q000276:immunology
chemical_list: D015798:HLA-DR1 Antigen; D010455:Peptides