Thymopoietin radioreceptor assay utilizing lectin-purified glycoprotein from a biologically responsive T cell line.
Radiolabeled thymopoietin that was biologically active and of high specific activity was prepared by a novel technique involving protection of free amino groups, selective excision of the protected N-terminal prolyl group with post-proline cleaving enzyme, reaction of the newly exposed alpha-amino group with a highly radioiodinated compound, and deprotection and purification of the polypeptide. Binding of this radiolabeled thymopoietin was not demonstrable by conventional techniques with cells, cell membranes, or solubilized cell membranes, apparently due to the presence of active proteases in these preparations. A glycoprotein with thymopoietin binding properties was prepared by lectin purification from the detergent-solubilized membranes of CEM cells, a human T cell line that responds to thymopoietin in vitro with increases in intracellular cyclic GMP. Presumably this procedure separated the thymopoietin binding protein from membrane proteases, thus permitting the development of a radioreceptor assay. Evidence is presented that the thymopoietin binding protein represents a thymopoietin receptor that is probably related to the mediation of immunoregulatory actions of thymopoietin on a subset of peripheral T cells.
D001667:Binding, Competitive; D002352:Carrier Proteins / Q000302:isolation & purification; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D002846:Chromatography, Affinity / Q000379:methods; D006152:Cyclic GMP / Q000378:metabolism; D006801:Humans; D037102:Lectins; D008565:Membrane Proteins / Q000302:isolation & purification*; D011485:Protein Binding; D011869:Radioligand Assay; D012995:Solubility; D013601:T-Lymphocytes / Q000032:analysis*; D013946:Thymopoietins / Q000032:analysis* / Q000302:isolation & purification* / Q000494:pharmacology; D013951:Thymus Hormones / Q000032:analysis* / Q000302:isolation & purification*
D002352:Carrier Proteins; D037102:Lectins; D008565:Membrane Proteins; D013946:Thymopoietins; D013951:Thymus Hormones; D006152:Cyclic GMP