Post-translational acquisition of insulin binding activity by the insulin proreceptor. Correlation to recognition by autoimmune antibody.
The post-translational acquisition of ligand binding activity by the insulin receptor was examined in 3T3-L1 adipocytes. In pulse-chase experiments with [35S] methionine, labeled receptor species were separated into "active" and "inactive" forms by affinity chromatography on insulin-agarose and then were characterized and quantitated. It was found that the newly translated high molecular weight proreceptor lacks the capacity to bind insulin. The acquisition of binding activity is relatively slow (t1/2 = 45 min) and occurs prior to conversion of the proreceptor to the mature alpha- and beta-subunits by proteolytic cleavage and maturation of its N-linked oligosaccharide chains (t1/2 = 3 h). Glycosylation appears to be required for this activation since the aglycoproreceptor, synthesized in the presence of tunicamycin, does not acquire insulin binding activity. However, once the proreceptor has acquired ligand binding activity, removal of its N-linked oligosaccharide chains with endoglycosidase H has no effect on the ability of the proreceptor to bind insulin. The modification of the proreceptor to bind insulin. The modification of the proreceptor that gives rise to insulin binding activity most likely involves a conformational change in the binding domain. A human autoimmune antibody that recognizes only the active insulin binding site does not interact with the inactive proreceptor, whereas a rabbit polyclonal antireceptor antibody recognizes all forms. Thus, the autoimmune antibody must recognize a new epitope created during conversion of the inactive proreceptor to the active form.
D000818:Animals; D000936:Antigen-Antibody Complex; D001323:Autoantibodies; D002478:Cells, Cultured; D007328:Insulin / Q000378:metabolism*; D011498:Protein Precursors / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011972:Receptor, Insulin / Q000235:genetics* / Q000276:immunology / Q000378:metabolism
D000936:Antigen-Antibody Complex; D001323:Autoantibodies; D007328:Insulin; D011498:Protein Precursors; C051512:insulin proreceptor; D011972:Receptor, Insulin