The Anti-angiogenic His/Pro-rich Fragment of Histidine-rich Glycoprotein Binds to Endothelial Cell Heparan Sulfate in a Zn2+-dependent Manner

• Published in: The Journal of biological chemistry 2006-04, Vol.281 (15), p.10298-10304
• Main Authors: Vanwildemeersch, Maarten, Olsson, Anna-Karin, Gottfridsson, Eva, Claesson-Welsh, Lena, Lindahl, Ulf, Spillmann, Dorothe
• Format: Article
• Language: English
• Subjects: Animals; Biotinylation; Cations; Cattle; Cell Line; Cell Membrane - metabolism; Cells, Cultured; Chemotaxis; Dose-Response Relationship, Drug; Endothelial Cells - metabolism; Endothelium, Vascular - cytology; Endothelium, Vascular - metabolism; Heparin - chemistry; Heparitin Sulfate - chemistry; Histidine - chemistry; Humans; Kinetics; Oligosaccharides - chemistry; Peptides - chemistry; Proline - chemistry; Protein Binding; Protein Structure, Tertiary; Recombinant Proteins - chemistry; Tissue Distribution; Zinc - chemistry
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M508483200

The plasma protein histidine-rich glycoprotein (HRGP), which has been identified as an angiogenesis inhibitor, binds to heparan sulfate (HS) in a Zn2+-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 Zn2+. We now show that HRGP330 binds heparin/HS with the same capacity as full-length HRGP, and the binding is Zn2+-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.