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Thromb Haemost 2003; 90(02): 309-316
DOI: 10.1160/TH03-01-0045
Platelets and Blood Cells
Schattauer GmbH

Affinity and kinetics of P-selectin binding to heparin

Jian-Guo Wang
1   Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China
,
Jian-Guo Geng
1   Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China
› Author AffiliationsFinancial support: This study was funded by grants from Chinese Academy of Sciences (KSCX2-2-02), National Science Foundation of China (39925015, 30130090 and 39970794), Special Funds for Major State Basic Research of China (G1999053907) and 973 Program (2002CB513000) from Chinese Ministry of Science and Technology.
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Publication History

Received 22 January 2003

Accepted after revision 13 May 2003

Publication Date:
06 December 2017 (online)

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Summary

P-selectin (CD62P), expressed on stimulated endothelial cells and activated platelets, reacts with P-selectin glycoprotein ligand-1 (PSGL-1, CD162) for leukocyte rolling. It also binds to heparin and heparan sulfate proteoglycans (HSPGs), which attenuates P-selectin mediated adhesions of leukocytes and cancer cells. Here we report that P-selectin mediated adhesion, but not rolling, of the HSPGs bearing human malignant melanoma A375 cells under shear stress. To understand its underlying molecular mechanism, we measured the biophysical properties of this interaction. Heparin inhibited the adhesion of A375 cells to immobilized P-selectin under flow (IC50 = 3 µM heparin) and neutralized the binding of P-selectin to A375 cells (IC50 = 4 µM heparin). Using surface plasmon resonance technique, we found that P-selectin bound to heparin with a dissociation constant (Kd ) of 115 ± 6 nM. The measured off rate (k off) was 3.15 ± 0.34 × 10-3 s-1 and the calculated on rate (k on) was 2.75 × 104 M-1 s-1. Taken together, our data suggest that the very slow k off and the reduced k on, but apparently not the Kd , are responsible for adhesion, but not rolling of A375 cells, to P-selectin under flow.

Keywords

P-selectin - heparin - heparan sulfate proteoglycans - affinity - kinetics
 

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