An investigation of the ligand binding properties and negative cooperativity of soluble insulin-like growth factor receptors.
To investigate the interaction of the insulin-like growth factor (IGF) ligands with the insulin-like growth factor type 1 receptor (IGF-1R), we have generated two soluble variants of the IGF-1R. We have recombinantly expressed the ectodomain of IGF-1R or fused this domain to the constant domain from the Fc fragment of mouse immunoglobulin. The ligand binding properties of these soluble IGF-1Rs for IGF-I and IGF-II were investigated using conventional ligand competition assays and BIAcore biosensor technology. In ligand competition assays, the soluble IGF-1Rs both bound IGF-I with similar affinities and a 5-fold lower affinity than that seen for the wild type receptor. In addition, both soluble receptors bound IGF-II with similar affinities to the wild type receptor. BIAcore analyses showed that both soluble IGF-1Rs exhibited similar ligand-specific association and dissociation rates for IGF-I and for IGF-II. The soluble IGF-1R proteins both exhibited negative cooperativity for IGF-I, IGF-II, and the 24-60 antibody, which binds to the IGF-1R cysteine-rich domain. We conclude that the addition of the self-associating Fc domain to the IGF-1R ectodomain does not affect ligand binding affinity, which is in contrast to the soluble ectodomain of the IR. This study highlights some significant differences in ligand binding modes between the IGF-1R and the insulin receptor, which may ultimately contribute to the different biological activities conferred by the two receptors.
Surinya
Katharina Helen
KH
School of Molecular and Biomedical Science, University of Adelaide, Adelaide 5005, Australia.
Forbes
Briony E
BE
Occhiodoro
Filomena
F
Booker
Grant W
GW
Francis
Geoffrey Leonard
GL
Siddle
Kenneth
K
Wallace
John C
JC
Cosgrove
Leah J
LJ
D000818:Animals; D006801:Humans; D007127:Immunoglobulin Constant Regions / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007334:Insulin-Like Growth Factor I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007335:Insulin-Like Growth Factor II / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008024:Ligands; D051379:Mice; D008956:Models, Chemical; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D017526:Receptor, IGF Type 1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020349:Surface Plasmon Resonance
D007127:Immunoglobulin Constant Regions; D008024:Ligands; D011993:Recombinant Fusion Proteins; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II; D017526:Receptor, IGF Type 1