Immunochemical analysis of the released Leu-2 (T8) molecule.
We recently have found that the human T cell antigen Leu-2 was specifically released from Leu-2-bearing cells. The preliminary study showed that the released Leu-2 (RLeu-2) from HPB-ALL cells was composed of a single polypeptide chain of 27,000 molecular weight (mol wt), which was smaller than the subunit of the homodimeric molecule found on the cell surface. In the present study, RLeu-2 was further characterized and compared with cellular Leu-2 (CLeu-2). Metabolically radiolabeled Leu-2 was released from HPB-ALL cells and this released Leu-2 molecule had a mol wt of 27,000. Cell surface radioiodinated HPB-ALL cells were found to release radioactive Leu-2 molecules and this antigen also had the same mol wt of 27,000. In both experiments, the CLeu-2 was reconfirmed to be composed of a 33,000-mol wt subunit under reducing conditions. These experiments establish that the 27,000-mol wt single polypeptide chain of Leu-2 released from the cell is derived directly from the homodimeric Leu-2 molecule on the cell surface, presumably by a specific proteolytic cleavage. Two-dimensional gel analysis showed that CLeu-2 exhibited extensive charge heterogeneity with predominantly basic isoelectric points, whereas RLeu-2 was a group of more acidic proteins with less charge heterogeneity. Although CLeu-2 and RLeu-2 showed several different immunochemical characteristics, the homology between these two antigens was confirmed by the following results: CLeu-2 and RLeu-2 were found to share at least three different antigenic determinants, Leu-2a and Leu-2b, and those which were detected by a polyvalent rabbit antiserum. Significant similarities between CLeu-2 and RLeu-2 were demonstrated by peptide mapping analysis of these antigens. Therefore, RLeu-2 appears to be the specific, physiological product of the CLeu-2 protein.
D000818:Animals; D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis; D006801:Humans; D051379:Mice; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism
D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface; D000939:Epitopes; D010446:Peptide Fragments