Analysis of the human insulin receptor.
The insulin derivative 4-azidosalicyloyl-[B1-biocytin-B2-lysine]insulin was used to photo-affinity-label the highly purified insulin receptor from human placenta. As shown by SDS-polyacrylamide gel electrophoresis, the 5 monoiodo isomers, with iodine in positions B1, B16, B26, A14 or A19, gave different labelling patterns. After complete tryptic digestion of the covalent receptor complex with 125I-Asa-[BctB1,LysB2]insulin, a stable fragment of 18 kDa was isolated, which was further purified by HPLC. This tryptic fragment of the intact receptor corresponds, according to HPLC, Tricin-SDS-PAGE and 2D-electrophoresis, to the similarly labelled sequenced domain of the receptor ectodomain (Fabry, M. et al. (1992) J. Biol. Chem. 267, 8950-8956). We thus conclude that insulin is bound to identical contact sites of native receptor and truncated ectodomain.
Fabry
M
M
Deutsches Wollforschungsinstitut, Technischen Hochschule Aachen.
Brandenburg
D
D
D000345:Affinity Labels / Q000378:metabolism*; D000818:Animals; D001360:Avidin / Q000378:metabolism; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D006801:Humans; D007328:Insulin / Q000031:analogs & derivatives / Q000378:metabolism; D007457:Iodine Radioisotopes; D046911:Macromolecular Substances; D010920:Placenta / Q000648:ultrastructure; D011247:Pregnancy; D011972:Receptor, Insulin / Q000378:metabolism*; D013552:Swine; D014357:Trypsin / Q000378:metabolism
D000345:Affinity Labels; D007328:Insulin; D007457:Iodine Radioisotopes; D046911:Macromolecular Substances; C074609:insulin, B1-(4-azidosalicyloyl)-(B1-biocytin,B2-lysine)-; D001360:Avidin; D011972:Receptor, Insulin; D014357:Trypsin