Competition for binding to insulin-like growth factor (IGF) binding protein-2, 3, 4, and 5 by the IGFs and IGF analogs.

The insulin-like growth factors (IGF) I and II bind to IGF binding proteins (BP) with high affinity. The affinity of each of the IGFs for individual BPs and the regions of the IGF-I molecule that are required for this high affinity binding have been defined only for IGFBP-1 and IGFBP-3. The present studies have determined the affinity of several IGF analogs (prepared using in vitro mutagenesis) for pure IGFBP-2, 3, 4, and 5. The results show IGFBP-2 binds these analogs in a manner similar to IGFBP-1. For example, a mutation in the A chain region (positions 49, 50, 51) or B chain (positions 3, 4) results in greater than 20-fold reduction in affinity for either IGFBP-1 or 2. In contrast, mutations in the A chain region have minimal effect on binding to IGFBP-3, whereas substitutions at the 3, 4, 15, 16 positions of the B chain reduce IGF-I affinity by at least 50-fold. At pH 7.4, binding of the analogs to IGFBP-4 is less affected by substitutions at the B chain 3, 4 positions compared to IGFBP-1, 2, and 3, but IGFBP-4 affinity for analogs containing the A chain substitutions is greatly reduced similarly to IGFBP-1 and 2. Binding to IGFBP-5 is greatly reduced by either A or B chain substitutions and most of the mutations result in greater than 100-fold reduction in affinity. Acidic pH 6.0 was associated with increased affinity of IGFBP-4 for the A chain containing mutants. The results indicate that only IGFBP-1 and 2 have nearly identical affinity for each of these analogs, whereas IGFBP-3, 4, and 5 have similarities and significant differences. The findings suggest that different binding proteins have differential structural requirements for optimal IGF-I binding.