title: Contributions of the N- and C-terminal domains of IGF binding protein-6 to IGF binding.
abstract: Insulin-like growth factors IGF-I and IGF -II are important mediators of growth. A family of six high affinity IGF binding proteins (IGFBPs) modulate IGF action. IGFBPs have three domains, of which the N- and C-domains are involved in high affinity IGF binding. IGFBP-6 is unique in its 20-100-fold IGF-II binding specificity over IGF-I. The aim of this study was to determine the contributions of the N- and C-domains of IGFBP-6 to its IGF binding properties. We confirmed that differential dissociation kinetics are responsible for the IGF-II binding preference of IGFBP-6. The N-domain has rapid association kinetics, similar to full-length IGFBP-6, but both IGF-I and -II dissociate rapidly from this domain, thereby reducing its binding affinity for IGF-II approximately 50-fold. However, the N-domain binds IGF-I and -II with similar affinities and it has a similar IGF-I binding affinity to full-length IGFBP-6. This suggests that the C-domain confers the IGF-II binding preference of IGFBP-6; indeed, IGF-I bound inconsistently with very low affinity to the C-domain. Coincubation studies showed that isolated N- and C-domains of IGFBP-6 do not strongly cooperate to enhance IGF binding. The results of the binding studies are supported by the effects of the IGFBP-6 domains on IGF-induced colon cancer cell proliferation; the N-domain inhibited IGF-II induced proliferation with approximately 20-fold lower potency than IGFBP-6 and it was equipotent in inhibiting IGF-I- and IGF-II-induced proliferation. Coincubation of C-domain had no additional effect on N-domain-induced inhibition of proliferation. In conclusion, both the N- and C-domains of IGFBP-6 are involved in IGF binding, the C-domain is responsible for the IGF-II binding preference of IGFBP-6 and intact IGFBP-6 is necessary for high affinity IGF binding.
authors.affiliation: University of Melbourne, Department of Medicine, Austin Hospital, Heidelberg 3084, Australia.
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Headey
authors.forename: Kerri S
authors.initials: KS
authors.lastname: Leeding
authors.forename: Raymond S
authors.initials: RS
authors.lastname: Norton
authors.forename: Leon A
authors.initials: LA
authors.lastname: Bach
mesh_terms: D015374:Biosensing Techniques; D049109:Cell Proliferation / Q000187:drug effects; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D006801:Humans; D018976:Insulin-Like Growth Factor Binding Protein 6 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000235:genetics / Q000378:metabolism*; D007335:Insulin-Like Growth Factor II / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D014407:Tumor Cells, Cultured
chemical_list: D018976:Insulin-Like Growth Factor Binding Protein 6; D010446:Peptide Fragments; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II