Structure function relationships for the IL 2 receptor system. III. Tac protein missing amino acids 102 to 173 (exon 4) is unable to bind IL 2: detection of spliced protein after L cell transfection.
High affinity receptors for interleukin 2 (IL 2) contain the Tac protein as one ligand-binding subunit. Localization of the IL 2-binding site on this molecule, as well as localization of the complementary site on IL 2, should provide insight into the design of IL 2 analogs. In this report, we examine the ability of normal and modified Tac protein to bind IL 2 and several antibodies that recognize the native Tac molecule. Using a transient L cell expression system, we have determined that transfection with cDNA-missing Tac exon 4 resulted in expression of spliced protein that had no measurable binding to IL 2 or the monoclonal anti-receptor antibodies, anti-Tac, and 7G7/B6. This protein was detected, however, by rabbit polyclonal antibodies prepared against synthetic Tac peptides. Thus, one or more amino acids encoded by exon 4 is important, either for direct ligand contact or for the proper folding of critical segments of the Tac molecule. In addition, insertion of stop codons at a unique restriction enzyme site near the beginning of exon 5 resulted in cellular secretion of truncated Tac molecules that were capable of binding IL 2, anti-Tac, and 7G7/B6. Amino acids encoded by exons 5 to 8 thus play no critical role in IL 2 binding. The ligand association demonstrated for truncated Tac protein produced by exons 2 to 4 should guide attempts to define the IL 2-binding segment of the Tac molecule.
D000595:Amino Acid Sequence; D000818:Animals; D000937:Antigen-Antibody Reactions; D000954:Antigens, Surface / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D004247:DNA / Q000235:genetics; D006801:Humans; D007376:Interleukin-2 / Q000378:metabolism*; D007739:L Cells; D051379:Mice; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D011971:Receptors, Immunologic / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015375:Receptors, Interleukin-2; D011994:Recombinant Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D014162:Transfection; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7
D000954:Antigens, Surface; D007376:Interleukin-2; D011971:Receptors, Immunologic; D015375:Receptors, Interleukin-2; D011994:Recombinant Proteins; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7; D004247:DNA