Structure of the IGF-binding domain of the insulin-like growth factor-binding protein-5 (IGFBP-5): Implications for IGF and IGF-I receptor interactions

Binding proteins for insulin-like growth factors (IGFs) IGF-I and IGF-II, k nown as IGFBPs, control the distribution, function and activity of IGFs in various cell tissues and body fluids. Insulin-like growth factor-binding pr otein-5 (IGFBP-5) is known to modulate the stimulatory effects of IGFs and is the major IGF-binding protein in bone tissue. We have expressed two N-te rminal fragments of IGFBP-5 in Escherichia coli; the first encodes the N-te rminal domain of the protein (residues 1-104) and the second, mini-IGFBP-5, comprises residues Ala40 to Ile92, We show that the entire IGFBP-5 protein contains only one high-affinity binding site for IGFs, located in mini-IGF BP-5. The solution structure of mini-IGFBP-5, determined by nuclear magneti c resonance spectroscopy, discloses a rigid, globular structure that consis ts of a centrally located three-stranded anti-parallel P-sheet, Its scaffol d is stabilized further by two inside packed disulfide bridges. The binding to IGFs, which is in the nanomolar range, involves conserved Leu and Val r esidues localized in a hydrophobic patch on the surface of the IGFBP-5 prot ein. Remarkably, the IGF-I receptor binding assays of IGFBP-5 showed that I GFBP-5 inhibits the binding of IGFs to the IGF-I receptor, resulting in red uction of receptor stimulation and autophosphorylation, Compared,vith the f ull-length IGFBP-5, the smaller N-terminal fragments were less efficient in hibitors of the IGF-I receptor binding of IGFs.