Structure and interactions of heparan sulfate proteoglycans from a mouse tumor basement membrane.
Various forms of heparan sulfate proteoglycan were solubilized from the mouse Engelbreth-Holm-Swarm (EHS) sarcoma by extraction with 0.5 M NaCl, collagenase digestion and extraction with 4 M guanidine. They could be separated into high (greater than or equal to 1.65 g/ml) and low (1.38 g/ml) buoyant density variants. The high-density form from the NaCl extract and collagenase digest had Mr = 130000 and So20,W = 4.5 S and contained 4-10% protein, indicating Mr = 5 000-12 000 for the protein core. This proteoglycan exhibited polydispersity as shown by rotary shadowing electron microscopy and ultracentrifugation. An average molecule consisted of four heparan sulfate chains (Mr = 29 000) each with a length of 32 +/- 10 nm. The low-density form (Mr about 400 000) could not be completely purified and contained about 50% protein. As shown by radioimmunoassay, the various proteoglycans shared similar protein cores. Labeling of the tumor in vivo or in vitro demonstrated preferential incorporation of radioactive sulfate in the high-density form. The high-density proteoglycan interacted in affinity chromatography by virtue of its heparan sulfate chains with laminin, fibronectin, the globular domain NC1 and the triple helix of collagen IV. These interactions were abolished at moderate concentrations of NaCl (0.1-0.2 M) and in the presence of heparin, chondroitin sulfate or dextran sulfate. Interactions with the globule NC1 could also be demonstrated by velocity band centrifugation in sucrose gradients and a binding constant of about 10(6) M-1 was derived.
D000818:Animals; D001485:Basement Membrane / Q000032:analysis; D002499:Centrifugation, Density Gradient; D011508:Chondroitin Sulfate Proteoglycans / Q000032:analysis*; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D006025:Glycosaminoglycans / Q000032:analysis*; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate / Q000032:analysis*; D051379:Mice; D008810:Mice, Inbred C57BL; D008854:Microscopy, Electron; D009994:Osmolar Concentration; D011509:Proteoglycans / Q000032:analysis*; D012513:Sarcoma, Experimental / Q000032:analysis*; D014461:Ultracentrifugation
D011508:Chondroitin Sulfate Proteoglycans; D006025:Glycosaminoglycans; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate; D011509:Proteoglycans