Purification of an insulin-like growth factor II receptor from rat chondrosarcoma cells.
An insulin-like growth factor II (IGF-II) receptor was purified from rat chondrosarcoma cells by Triton X-100 solubilization of a 100,000 X g membrane preparation and affinity chromatography on a multiplication-stimulating activity (MSA)-Sepharose column. Analysis of the purified receptor by sodium dodecyl sulfate-gel electrophoresis and silver staining showed a major band of Mr = 210,000 (Mr = 250,000 after reduction with dithiothreitol). When 125I-MSA was chemically cross-linked to the purified receptor and analyzed by sodium dodecyl sulfate-gel electrophoresis (with and without dithiothreitol) and autoradiography, the radioactive bands coincided with the Mr = 210,000 and 250,000 bands identified by silver staining. The purified receptor also appeared to contain an Mr = less than 68,000 species identified by silver staining in addition to the Mr = 250,000 binding component. IGF-I, IGF-II, and MSA-II inhibited binding of 125I-MSA to the purified receptor with the same relative potency as for binding to the intact chondrosarcoma cell (IGF-II greater than MSA-II greater than IGF-I), and insulin did not inhibit binding. The association constant (K alpha) for MSA-II binding to the purified receptor was 2 X 10(9) M-1. The purified receptor bound to concanavalin A-Sepharose and wheat germ lectin-Sepharose columns and was eluted with alpha-methyl-D-mannoside and N-acetyl-D-glucosamine, respectively, showing that the receptor is a glycoprotein.
D000818:Animals; D002846:Chromatography, Affinity; D003432:Cross-Linking Reagents / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D007335:Insulin-Like Growth Factor II; D008970:Molecular Weight; D012516:Osteosarcoma / Q000032:analysis*; D010455:Peptides / Q000378:metabolism; D051381:Rats; D011956:Receptors, Cell Surface / Q000302:isolation & purification* / Q000378:metabolism; D017451:Receptors, Somatomedin
D003432:Cross-Linking Reagents; D010455:Peptides; D011956:Receptors, Cell Surface; D017451:Receptors, Somatomedin; D007335:Insulin-Like Growth Factor II