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Purification of an insulin-like growth factor II receptor from rat chondrosarcoma cells.

1983; Elsevier BV; Volume: 258; Issue: 15 Linguagem: Inglês

10.1016/s0021-9258(17)44625-4

1083-351X

Gilbert P. August, S. Peter Nissley, Masato Kasuga, L Lee, Lawrence A. Greenstein, Matthew M. Rechler,

Cancer, Hypoxia, and Metabolism

An insulin-like growth factor I1 (IGF-11) 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 M , = 210,000 (Mr = 250,000 after reduction with dithiothreitol).When '"I-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 M, = 210,000 and 250,000 bands identified by silver staining.The purified receptor also appeared to contain an M. = ~6 8 , 0 0 0 species identified by silver staining in addition to the M, = 250,000 binding component.IGF-I, IGF-11, and MSA-I1 inhibited binding of '"I-MSA to the purified receptor with the same relative potency as for binding to the intact chondrosarcoma cell (IGF-I1 > MSA-I1 > IGF-I), and insulin did not inhibit binding.The association constant ( I C 4 ) for MSA-I1 binding to the purified receptor was 2 X lo9 M " .The purified receptor bound to concanavalin A-Sepharose and wheat germ lectin-Sepharose columns and was eluted with a-methyl-D-mannoside and N-acetyl-D-ghcosamine, respectively, showing that the receptor is a glycoprotein.