Insulin-like growth factor-binding protein 5 complexes with the acid-labile subunit. Role of the carboxyl-terminal domain.
We have recently shown that insulin-like growth factor (IGF)-binding protein 5 forms ternary complexes with IGF-I or IGF-II and the acid-labile subunit (ALS) (Twigg, S. M., and Baxter, R. C. (1998) J. Biol. Chem. 273, 6074-6079). Because IGF-binding protein 3 (IGFBP-3) binds to ALS through its basic carboxyl-terminal domain, we tested whether a homologous region present in IGFBP-5 is involved in IGFBP-5 binding to ALS. Chimeric peptides were generated by carboxyl-terminal domain interchange between recombinant human IGF-BP-5 and IGFBP-6, producing two IGFBP peptides designated 5-5-6 and 6-6-5. Determined by immunoprecipitation and by Superose chromatography, 6-6-5 formed ternary complexes, albeit less potently than IGF-BP-5. In contrast, 5-5-6, like IGFBP-6, did not form ternary complexes by these methods. Whereas 6-6-5, like IGFBP-6, had a marked preference for binary complex formation with IGF-II rather than IGF-I, it formed ternary complexes more efficiently with IGF-I, like IGF-BP-5. The glycosaminoglycans heparin and heparan sulfate bind to IGFBP-5 through its basic carboxyl-terminal domain. At high concentrations, these glycosaminoglycans inhibited ALS binding to binary complexed IGF-BP-5. In addition, in the absence of IGFs, IGFBP-5, a synthetic peptide representing the basic carboxyl-terminal sequence IGFBP-5(201-218), and the corresponding IGFBP-3 basic sequence IGFBP-3(215-232), competed weakly for ALS binding to covalent IGF-IGFBP-5 complex, as did a random-sequence synthetic peptide with the same composition as IGFBP-5(201-218). These findings are consistent with the basic carboxyl-terminal domain on IGFBP-5 being the principal site in IGFBP-5 that binds to ALS.
Twigg
S M
SM
Kolling Institute of Medical Research, University of Sydney, Royal North Shore Hospital, St. Leonards, New South Wales 2065, Australia.
Kiefer
M C
MC
Zapf
J
J
Baxter
R C
RC
D000595:Amino Acid Sequence; D001667:Binding, Competitive; D006025:Glycosaminoglycans / Q000378:metabolism; D006801:Humans; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism
D006025:Glycosaminoglycans; D018975:Insulin-Like Growth Factor Binding Protein 5; D011993:Recombinant Fusion Proteins