The heparin binding domain of insulin-like growth factor binding protein (IGFBP)-3 increases susceptibility of IGFBP-3 to proteolysis

IGFBP-3 proteolysis clears IGFBP-3 from body fluids and increases IGF bioav ailability. As shown here, native human IGFBP-3 was cleaved by proteases in media conditioned by hamster and insect cells. This proteolysis was less p ronounced for IGFBP-3 containing a mutated heparin binding domain, and was prevented by purifying IGFBP-3 on an IGF-l affinity column in the presence of 2M sodium chloride, suggesting that the responsible protease(s) binds th e IGFBP-3 heparin binding domain. To determine if any human proteases act t his way, we first studied plasma prekallikrein since it can copurify with I GFBP-3, and found: 1) [(125)]IGFBP-3 binds to prekallikrein immobilized eit her on nitrocellulose or on immunocapture plates; 2) the IGFBPS heparin bin ding domain participates in forming the IGFBP-3/prekallikrein complex; 3) t he binary IGFBP-3/prekallikrein complex can bind IGF-I to form a ternary co mplex; and 4) activation of prekallikrein to alpha-kallikrein by Factor XII a resulted in proteolysis of bound IGFBP-3. This work suggests: 1) cleavage of IGFBP-3 by a protease may be aided by the ability of the protease zymog en to directly bind the IGFBP-3 heparin binding domain; and 2) direct bindi ng of protease zymogens to IGFBP-3 may explain some instances where IGFBP-3 is preferentially proteolyzed in the presence of other IGFBPs.