title: Detection of a new hormone contact site within the insulin receptor ectodomain by the use of a novel photoreactive insulin.
abstract: We have used a preparation of soluble human insulin receptor ectodomain and a novel photoreactive, biotinylated derivative of insulin (4-azidosalicyloyl(B1-biocytinyl-B2-lysine)-insulin) to identify a new hormone contact site within the extracellular domain of the insulin receptor. The ectodomain was photoaffinity-labeled and digested to completion with trypsin, and the resulting tryptic fragment was purified by either HPLC or by streptavidin-affinity chromatography. The amino terminus of the fragment was identified as Gly390 within the alpha-subunit. These results suggest that residues that are carboxyl-terminal to the cysteine-rich domain, in addition to previously identified regions within the amino terminus of the alpha-subunit, contribute to the insulin binding site. The implications of these results for the de novo folding of the insulin receptor to constitute the hormone binding site are discussed.
authors.affiliation: Deutsches Wollforschungsinstitut, Technischen Hochschule Aachen, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Fabry
authors.forename: E
authors.initials: E
authors.lastname: Schaefer
authors.forename: L
authors.initials: L
authors.lastname: Ellis
authors.forename: E
authors.initials: E
authors.lastname: Kojro
authors.forename: F
authors.initials: F
authors.lastname: Fahrenholz
authors.forename: D
authors.initials: D
authors.lastname: Brandenburg
mesh_terms: D000345:Affinity Labels; D000595:Amino Acid Sequence; D000818:Animals; D001386:Azides; D001665:Binding Sites; D016466:CHO Cells; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006868:Hydrolysis; D007328:Insulin / Q000031:analogs & derivatives / Q000378:metabolism*; D008969:Molecular Sequence Data; D010777:Photochemistry; D011972:Receptor, Insulin / Q000235:genetics / Q000378:metabolism*; D012459:Salicylates; D014357:Trypsin / Q000378:metabolism
chemical_list: D000345:Affinity Labels; D001386:Azides; D007328:Insulin; D012459:Salicylates; C063873:4-azidosalicylic acid; D011972:Receptor, Insulin; D014357:Trypsin