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J Biol Chem, Vol. 274, Issue 18, 12376-12382, April 30, 1999

Characterization of a Neutrophil Cell Surface Glycosaminoglycan That Mediates Binding of Platelet Factor 4

Frank PetersenDagger , Ernst Brandt, Ulf LindahlDagger , and Dorothe SpillmannDagger

From the Dagger  Department of Medical Biochemistry and Microbiology, Uppsala University, Biomedical Center, Box 575, S-75123 Uppsala, Sweden and the  Department of Immunology and Cell Biology, Research Center Borstel, Parkallee 22, D-23845 Borstel, Germany

Platelet factor 4 (PF-4) is a platelet-derived alpha -chemokine that binds to and activates human neutrophils to undergo specific functions like exocytosis or adhesion. PF-4 binding has been shown to be independent of interleukin-8 receptors and could be inhibited by soluble chondroitin sulfate type glycosaminoglycans or by pretreatment of cells with chondroitinase ABC. Here we present evidence that surface-expressed neutrophil glycosaminoglycans are of chondroitin sulfate type and that this species binds to the tetrameric form of PF-4. The glycosaminoglycans consist of a single type of chain with an average molecular mass of ~23 kDa and are composed of ~85-90% chondroitin 4-sulfate disaccharide units type CSA (right-arrow4GlcAbeta 1right-arrow3GalNAc(4-O-sulfate)beta 1right-arrow) and of ~10-15% di-O-sulfated disaccharide units. A major part of these di-O-sulfated disaccharide units are CSE units (right-arrow4GlcAbeta 1right-arrow3GalNAc(4,6-O-sulfate)beta 1right-arrow). Binding studies revealed that the interaction of chondroitin sulfate with PF-4 required at least 20 monosaccharide units for significant binding. The di-O-sulfated disaccharide units in neutrophil glycosaminoglycans clearly promoted the affinity to PF-4, which showed a Kd ~ 0.8 µM, as the affinities of bovine cartilage chondroitin sulfate A, porcine skin dermatan sulfate, or bovine cartilage chondroitin sulfate C, all consisting exclusively of monosulfated disaccharide units, were found to be 3-5-fold lower. Taken together, our data indicate that chondroitin sulfate chains function as physiologically relevant binding sites for PF-4 on neutrophils and that the affinity of these chains for PF-4 is controlled by their degree of sulfation.


Copyright © 1999 by The American Society for Biochemistry and Molecular Biology, Inc.
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