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 bioavailability. As shown here, native human IGFBP-3 was cleaved by proteases in media conditioned by hamster and insect cells. This proteolysis was less pronounced for IGFBP-3 containing a mutated heparin binding domain, and was prevented by purifying IGFBP-3 on an IGF-I affinity column in the presence of 2 M sodium chloride, suggesting that the responsible protease(s) binds the IGFBP-3 heparin binding domain. To determine if any human proteases act this way, we first studied plasma prekallikrein since it can copurify with IGFBP-3, and found: 1) [125]IGFBP-3 binds to prekallikrein immobilized either on nitrocellulose or on immunocapture plates; 2) the IGFBP-3 heparin binding domain participates in forming the IGFBP-3/prekallikrein complex; 3) the binary IGFBP-3/prekallikrein complex can bind IGF-I to form a ternary complex; and 4) activation of prekallikrein to alpha-kallikrein by Factor XIIa 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 zymogen to directly bind the IGFBP-3 heparin binding domain; and 2) direct binding of protease zymogens to IGFBP-3 may explain some instances where IGFBP-3 is preferentially proteolyzed in the presence of other IGFBPs.
Durham
S K
SK
Department of Pediatrics, Baylor College of Medicine, Houston, TX, USA.
Suwanichkul
A
A
Hayes
J D
JD
Herington
A C
AC
Powell
D R
DR
Campbell
P G
PG
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005174:Factor XII / Q000378:metabolism; D015870:Gene Expression / Q000502:physiology; D006493:Heparin / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D018970:Insulin-Like Growth Factor Binding Protein 1 / Q000235:genetics; D018971:Insulin-Like Growth Factor Binding Protein 2 / Q000235:genetics; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000097:blood / Q000737:chemistry / Q000235:genetics; D007334:Insulin-Like Growth Factor I / Q000235:genetics; D007335:Insulin-Like Growth Factor II / Q000235:genetics; D007457:Iodine Radioisotopes; D007610:Kallikreins / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis / Q000502:physiology; D015345:Oligonucleotide Probes; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary
D018970:Insulin-Like Growth Factor Binding Protein 1; D018971:Insulin-Like Growth Factor Binding Protein 2; D018972:Insulin-Like Growth Factor Binding Protein 3; D007457:Iodine Radioisotopes; D015345:Oligonucleotide Probes; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II; D005174:Factor XII; D006493:Heparin; D007610:Kallikreins