The anti-angiogenic His/Pro-rich fragment of histidine-rich glycoprotein binds to endothelial cell heparan sulfate in a Zn2+-dependent manner.
The plasma protein histidine-rich glycoprotein (HRGP), which has been identified as an angiogenesis inhibitor, binds to heparan sulfate (HS) in a Zn(2+)-dependent manner. We wished to test whether this interaction is mechanistically important in mediation of the anti-angiogenic effect of HRGP. Inhibition of angiogenesis by HRGP is exerted through its central His/Pro-rich domain, which is proteolytically released. A 35-amino-acid residue synthetic peptide, HRGP330, derived from the His/Pro-rich domain retains the inhibitory effect on blood vessel formation in vitro and in vivo, an effect dependent on the presence of Zn(2+). We now show that HRGP330 binds heparin/HS with the same capacity as full-length HRGP, and the binding is Zn(2+)-dependent. Peptides derived from the His/Pro-rich domain of HRGP downstream of HRGP330 fail to inhibit endothelial cell migration and display a significantly reduced heparin-binding capacity. An even shorter peptide, HRGP335, covering a 26-amino-acid sequence within HRGP330 retains full heparin/HS-binding capacity. Characterization of the HS interaction shows that there is a tissue-specific HS pattern recognized by HRGP335 and that the minimal length of heparin/HS required for binding to HRGP335 is an 8-mer oligosaccharide. Saturation of the HS binding sites in HRGP330 by pre-incubation with heparin abrogates the HRGP330-induced rearrangement of endothelial cell focal adhesions, suggesting that interaction with cell surface HS is needed for HRGP330 to exert its anti-angiogenic effect.
Vanwildemeersch
Maarten
M
Department of Medical Biochemistry and Microbiology, Uppsala University, SE-75123 Uppsala, Sweden.
Olsson
Anna-Karin
AK
Gottfridsson
Eva
E
Claesson-Welsh
Lena
L
Lindahl
Ulf
U
Spillmann
Dorothe
D
D000818:Animals; D019921:Biotinylation; D002412:Cations; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002633:Chemotaxis; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D006493:Heparin / Q000737:chemistry; D006497:Heparan Sulfate / Q000737:chemistry*; D006639:Histidine / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D009844:Oligosaccharides / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D011392:Proline / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D014018:Tissue Distribution; D015032:Zinc / Q000737:chemistry*
D002412:Cations; D006493:Heparin; D006497:Heparan Sulfate; D006639:Histidine; D009844:Oligosaccharides; D010455:Peptides; D011392:Proline; D011994:Recombinant Proteins; D015032:Zinc