title: Insulin and IGF binding by IGFBP-3 fragments derived from proteolysis, baculovirus expression and normal human urine.
abstract: Recombinant human IGFBP-3 was proteolysed with different concentrations of plasmin for various periods of time. The major IGFBP-3 fragment resulting from this digestion migrated at ca. 15 kDa in nonreducing SDS-PAGE. Following the identification of this fragment as an N-terminal IGFBP-3 fragment, by use of N-terminus-specific monoclonal antibody and amino acid sequence analysis, we constructed and expressed a similar fragment in a baculovirus expression system. The fragments resulting from plasmin digestion, as well as the baculovirus-expressed recombinant human IGFBP-3(1-97), retain weak IGF binding and show specific insulin binding on cross-linking and western ligand blot. RhIGFBP-3(1-97) can inhibit insulin receptor autophosphorylation in insulin receptor-overexpressing NIH 3T3 cells. Insulin and IGF binding to IGFBP-3 fragments could be further demonstrated in normal urine. These data indicate the physiological significance of IGFBP-3 fragments derived from proteolysis in vivo.
authors.affiliation: Dept. Pediatrics NRC5, Oregon Health Sciences University, Portland 97201-3042, USA.
authors.forename: P
authors.initials: P
authors.lastname: Vorwerk
authors.forename: Y
authors.initials: Y
authors.lastname: Yamanaka
authors.forename: A
authors.initials: A
authors.lastname: Spagnoli
authors.forename: Y
authors.initials: Y
authors.lastname: Oh
authors.forename: R G
authors.initials: RG
authors.lastname: Rosenfeld
mesh_terms: D016367:Baculoviridae / Q000235:genetics*; D002675:Child, Preschool; D005260:Female; D005341:Fibrinolysin / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D007328:Insulin / Q000378:metabolism*; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000737:chemistry* / Q000652:urine; D010446:Peptide Fragments / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011994:Recombinant Proteins / Q000096:biosynthesis; D012016:Reference Values; D013002:Somatomedins / Q000378:metabolism*
chemical_list: D007328:Insulin; D018972:Insulin-Like Growth Factor Binding Protein 3; D010446:Peptide Fragments; D011994:Recombinant Proteins; D013002:Somatomedins; D005341:Fibrinolysin