Antigen conformation determines processing requirements for T-cell activation.
We studied the difference in requirements for processing and presentation to a single T-cell clone of four different forms of the same epitope of sperm whale myoglobin--namely, on the native protein, on two conformationally altered forms of the protein, or as a 22-residue antigenic peptide fragment. The T-cell clone was I-Ed-restricted and specific for an epitope on the CNBr fragment 132-153 involving Lys-140. As inhibitors of macrophage processing of antigen, we used several agents that inhibit lysosomal function: the weak bases chloroquine and NH4Cl, the cationic ionophore monensin, and the competitive protease inhibitor leupeptin. When these agents were used to inhibit processing of antigen by presenting cells and then washed out before T cells were added to culture, they inhibited the presentation of native antigen but not of fragment 132-153. To our surprise, the intact but denatured form, S-methylmyoglobin, behaved like the fragment not like the native protein. Apomyoglobin was intermediate in susceptibility to inhibition. Thus, native myoglobin requires a processing step that appears to involve lysosomal proteolysis, which is not required by fragment 132-153 or the denatured unfolded forms. For an antigen the size of myoglobin (Mr 17,800), it appears that unfolding of the native conformation, rather than further reduction in size, is the critical parameter determining the need for processing. Since a major difference between native myoglobin and the other forms is the greater accessibility in the latter of sites, such as hydrophobic residues, buried in the native protein, we propose that processing may be necessary to expose these sites, perhaps for interaction with the cell membrane or the Ia of the antigen-presenting cell.
D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D000941:Antigens / Q000276:immunology*; D002738:Chloroquine / Q000494:pharmacology; D002999:Clone Cells / Q000276:immunology; D007976:Leupeptins / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects; D008247:Lysosomes / Q000187:drug effects / Q000201:enzymology; D051379:Mice; D008985:Monensin / Q000494:pharmacology; D009211:Myoglobin / Q000276:immunology* / Q000378:metabolism; D010446:Peptide Fragments / Q000276:immunology; D011487:Protein Conformation; D011489:Protein Denaturation; D013329:Structure-Activity Relationship; D013601:T-Lymphocytes / Q000276:immunology*
D000941:Antigens; D007976:Leupeptins; D009211:Myoglobin; D010446:Peptide Fragments; C043441:S-methylmyoglobin; D000643:Ammonium Chloride; D002738:Chloroquine; D008985:Monensin; C032854:leupeptin