Covalent cross-linking of insulin-like growth factor-1 to a specific inhibitor from human serum.

Previous studies have shown that a specific inhibitor of insulin-like growth factor (IGF) action in vitro can be isolated from normal human serum and subsequently partially purified on an IGF-affinity column. The ability of the inhibitor to bind the IGFs has now been confirmed directly using covalent cross-linking techniques. When 125I-IGF-1 was cross-linked to inhibitor using disuccinimidyl suberate, five specifically labelled bands were seen on SDS-PAGE and autoradiography. Two bands (MW 21.5 K and 25.5 K) were intensely labelled, whilst the remaining three (MW 37 K, 34K and 18 K) appeared as minor bands only. Inhibitor bioactivity, following further analysis by hydrophobic interaction chromatography or Con A-Sepharose affinity chromatography, was always associated with the presence of the 21.5 K and/or 25.5 K bands. These data describe, for the first time, the structural nature of the IGF inhibitor protein and raise important questions regarding the relationship of the inhibitor to the primary IGF-binding subunit of the native high MW IGF carrier protein of serum.