Purified preparations of the amniotic fluid-derived insulin-like growth factor-binding protein contain multimeric forms that are biologically active.
The insulin-like growth factors (IGFs) appear to be secreted into interstitial fluids by many cell types, along with specific, high affinity binding proteins (IGF-BPs). These proteins, therefore, have the potential to bind IGF-I and -II and modify their ability to interact with specific cell surface receptors In these studies we report the detection of high mol wt, multimeric forms of one form of IGF-BP that has been purified from human amniotic fluid. The multimeric forms, which are either not or barely detectable in native amniotic fluid, are the result of intermolecular disulfide bond formation and can be reduced to a monomeric form by exposure to dithiothreitol. After reduction, the multimers are reduced to either monomeric or dimeric forms, as detected by Western blotting. The multimers can be separated from monomeric and dimeric forms by gel filtration chromatography. The purified multimers were fully biologically active in potentiating the effect of IGF-I on porcine aortic smooth muscle cell DNA synthesis. The monomeric form was also bioactive. No significant differences in the affinity of the monomeric and multimeric forms for IGF-I or -II could be detected. In summary, multimeric forms of this form of IGF-BP are detected during purification. The formation of these multimers is through intermolecular disulfide bonds and does not disrupt IGF binding or potentiation of the cellular growth response to IGF-I. These findings indicate that these higher mol wt forms may be fully active in biological test systems.
Busby
W H
WH
Department of Medicine, University of North Carolina School of Medicine, Chapel Hill 27599.
Hossenlopp
P
P
Binoux
M
M
Clemmons
D R
DR
D000653:Amniotic Fluid / Q000032:analysis*; D000818:Animals; D002352:Carrier Proteins / Q000032:analysis* / Q000302:isolation & purification / Q000378:metabolism; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D018969:Insulin-Like Growth Factor Binding Proteins; D007334:Insulin-Like Growth Factor I / Q000378:metabolism; D007335:Insulin-Like Growth Factor II / Q000378:metabolism; D009130:Muscle, Smooth / Q000166:cytology / Q000378:metabolism; D013552:Swine
D002352:Carrier Proteins; D018969:Insulin-Like Growth Factor Binding Proteins; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II