OH. treatment of tetanus toxin reduces its susceptibility to limited proteolysis with more efficient presentation to specific T cells.
At inflammatory sites, before their processing, antigens are exposed to oxygen free radicals released by activated cells. The effect of hydroxyl radicals (OH.) on the structure of a protein antigen, tetanus toxin (TT) was investigated, as well as the consequences on processing and presentation. A chemical system composed of Fe-EDTA, ascorbate and H2O2 was used to produce physiological amounts of OH. radicals. TT exposed to OH. radicals presented a marked decrease of its intrinsic fluorescence with a concomitant increase of the content of bityrosine, but no fragmentation of the protein was detected by SDS-PAGE. Processing of the modified TT was analysed, by incubating TT at acidic pH with fractions enriched in plasma membranes and lysosomes obtained from a lymphoblastoid cell line (LCL). Proteolysis of OH.-treated TT was less important than proteolysis of native TT, especially upon prolonged incubations. Oxidized TT presented by LCL cells induced a greater proliferation of three different TT specific T cell clones, compared to native TT. When proteolytic digests of TT were presented by fixed LCL cells to a homologous T cell line, the proliferative response obtained in the presence of digests of OH.-treated TT was sustained, even in the case of prolonged proteolysis, whereas the response to digests of native TT fell rapidly. The relative resistance of OH.-treated TT to proteolysis appears thus responsible for its greater presentation to specific T cells, probably by protecting epitopes.
Pernollet
M
M
Département de Biologie Moléculaire et Structurale, Laboratoire d'Immunochimie, INSERM Unité 238, Grenoble, France.
Villiers
C
C
Gabert
F
F
Drouet
C
C
Colomb
M
M
D000818:Animals; D000938:Antigen-Presenting Cells / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology*; D002478:Cells, Cultured; D002999:Clone Cells / Q000276:immunology; D010450:Endopeptidases / Q000378:metabolism; D006801:Humans; D017665:Hydroxyl Radical / Q000494:pharmacology*; D008213:Lymphocyte Activation / Q000187:drug effects; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000032:analysis / Q000276:immunology* / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D013347:Subcellular Fractions / Q000378:metabolism; D013601:T-Lymphocytes / Q000276:immunology; D013744:Tetanus Toxin / Q000276:immunology* / Q000378:metabolism* / Q000494:pharmacology; D014443:Tyrosine / Q000031:analogs & derivatives / Q000032:analysis
D010446:Peptide Fragments; D013744:Tetanus Toxin; D017665:Hydroxyl Radical; D014443:Tyrosine; C007543:dityrosine; D010450:Endopeptidases