Purification, cDNA structure and biological significance of a single insulin-like growth factor-binding domain protein (SIBD-1) identified in the hemocytes of the spider Cupiennius salei

Insulin-like growth factor binding proteins (IGFBPs) -3 and -5 are known to interact with various components of the extracellular matrix (ECM; e.g. heparin and heparan sulphate) and this interaction is believed to affect the affinity of both IGFBP species for their cognate ligands – IGF-I and -II. There is little detail on the nature of the molecular complex formed between ECM components, IGFBPs and IGFs although the glycosaminoglycan (GAG) heparin has been reported to reduce the affinity of IGFBP-5 for IGF-I. In order to investigate this phenomenon further, we have undertaken an extensive surface plasmon resonance based biosensor study to report the affinity of IGFBP-3 and -5 for binding heparin (22 and 7 nM respectively). We have also shown that pre-complexation of IGFBP with IGF-I and -II inhibits the subsequent association of IGFBP with heparin and conversely that heparin complexation of IGFBP-3 and -5 inhibits IGFBP binding to biosensor surfaces containing immobilised IGF-I. In addition we have used both IGF-I and heparin coated biosensor surfaces in an attempt to build ternary IGF–IGFBP–heparin complexes in order to gain some insight into the nature of inhibition by heparin of IGFI–IGFBP complex formation. Our data lead us to conclude that the inhibition by heparin is partly competitive in nature, and that ternary complexes of IGF–IGFBP–heparin are either unable to form, or only form unstable transient complexes. The potential biological significance of our data is highlighted by the demonstration that IGF-I and IGF-II can displace endogenous IGFBP-5 from monolayer cultures of the mouse mammary epithelial cell line HC11.

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Two site-specific radioimmunoassays which demonstrate the presence of proteolytically modified insulin-like growth factor-binding protein-3 in the circulation

Journal of Endocrinology ◽

10.1677/joe.0.1370321 ◽

1993 ◽

Vol 137 (2) ◽

pp. 321-328 ◽

Cited By ~ 8

Author(s):

S. C. Cwyfan Hughes ◽

J. A. H. Wass ◽

J. M. P. Holly

Keyword(s):

Growth Factor ◽

Binding Protein ◽

Antigenic Site ◽

Binding Domain ◽

Insulin Like Growth Factor ◽

Factor Binding ◽

Sds Page ◽

Igf Binding ◽

Harsh Conditions ◽

Igfbp 3

ABSTRACT The presence of proteolytic activity in the circulation directed against insulin-like growth factor binding protein-3 (IGFBP-3) was originally described in pregnancy but has subsequently been described in a number of catabolic and other pathological conditions. However, detection of this proteolytically modified IGFBP-3 has been demonstrated, in most cases, following separation using SDS-PAGE and this has led to speculation that its occurrence may be an artefact of the harsh conditions employed. Using two sitespecific antisera, one of which recognizes as its antigenic site a region of IGFBP-3 which is close to that of the IGF-binding domain, we have developed two radioimmunoassays which, when compared, can reveal alterations in the IGF-binding domain of IGFBP-3. The presence of IGFBP-3 modified by the circulating protease is denoted in this system by a divergence in the IGFBP-3 levels observed; this divergence has been shown to coincide with protease activity as determined by Western ligand blotting. The use of these assays has confirmed that the IGF-binding site of IGFBP-3 undergoes proteolytic modification in the circulation during a number of pathologies. Journal of Endocrinology (1993) 137, 321–328

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The heparin-binding domain of IGFBP-2 has insulin-like growth factor binding-independent biologic activity in the growing skeleton.

Journal of Biological Chemistry ◽

10.1074/jbc.a110.193334 ◽

2011 ◽

Vol 286 (50) ◽

pp. 43588.2-43588

Author(s):

Masanobu Kawai ◽

Anne C. Breggia ◽

Victoria E. DeMambro ◽

Xinchun Shen ◽

Ernesto Canalis ◽

...

Keyword(s):

Growth Factor ◽

Binding Domain ◽

Insulin Like Growth Factor ◽

Heparin Binding ◽

Factor Binding ◽

Biologic Activity ◽

Heparin Binding Domain

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Plasmin degradation of insulin-like growth factor-binding protein-5 (IGFBP-5): regulation by IGFBP-5-(201—218)

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1997.273.5.e996 ◽

1997 ◽

Vol 273 (5) ◽

pp. E996-E1004 ◽

Cited By ~ 4

Author(s):

Phil G. Campbell ◽

Dennis L. Andress

Keyword(s):

Growth Factor ◽

Solid Phase ◽

Binding Protein ◽

Competitive Inhibitor ◽

Binding Domain ◽

Tissue Type ◽

Insulin Like Growth Factor ◽

Heparin Binding ◽

Factor Binding ◽

Heparin Binding Domain

Using the major bone insulin-like growth factor-binding protein (IGFBP) IGFBP-5, we took a mechanistic approach in evaluating the role of the heparin-binding domain of IGFBP-5 in regulating plasmin (Pm) proteolysis of IGFBP-5. Using synthetic IGFBP-5 peptide fragments, we determined that the heparin-binding domain, IGFBP-5-(208—218), inhibits Pm proteolysis of intact IGFBP-5. The mechanism of action of IGFBP-5-(201—218) was by inhibiting Pm binding to substrate IGFBP-5. IGFBP-5-(201—218) action was independent of site of proteolysis, fluid, or solid phase interaction. In addition, IGFBP-5-(201—218) was found to inhibit plasminogen (Pg) activation to Pm. IGFBP-5-(201—218) did not directly inhibit the activity of Pm, urokinase Pg activator (PA), or tissue-type PA but acted as a competitive inhibitor of Pg activation by PA, which is in contrast to the stimulating effect of heparin on Pg activation. These data indicate that the heparin-binding domain contains the serine protease (Pg-to-Pm) binding site region of IGFBP-5, and that this region, which is presumed to represent a Pm-induced proteolytic product of IGFBP-5, is capable of regulating Pm action.

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