Heparin proteoglycans synthesized by mouse mastocytoma contain chondroitin sulphate.

• Proteoglycans (PGs), biosynthetically labelled with [S-35]sulphate, were isolated from mouse mastocytoma tissue. Chromatography on antithrombin (AT)-Sepharose resulted in the separation of the S-35-labelled PGs into three fractions: PGs with no affinity for the gel (NA-PGs), PGs with low affinity (LA-PGs), and PGs with high affinity (HA-PGs) for antithrombin. Whereas NA-PGs contained almost exclusively chondroitin sulphate (CS), the AT-binding PGs contained 80-85% heparin and 15-20% CS. [S-35]CS-containing macromolecules obtained from the HA-PG fraction after removal of the heparin polysaccharide chains were rechromatographed on AT-Sepharose. A majority of these S-35-labelled macromolecules no longer showed affinity for AT. These experiments indicate that the [S-35]CS recovered in the AT-binding PGs is present in hybrid PGs. Polysaccharide chain-length determination demonstrated that the heparin chains were somewhat larger (M(r) similar to 30000) than the CS chains in the NA-PGs (M(r) similar to 25000). CS chains in the hybrid PGs were slightly smaller (M(r) similar to 20000). Characterization of the sulphated CS disaccharides from NA- and HA-PGs showed that they contained similar amounts (20 %) of disulphated disaccharides of [GlcA-GalNAc(4,6-di-OSO3)] type. The monosulphated CS-disaccharides were O-sulphated at C-4 of the galactosamine units. Analysis by gel chromatography of the [S-35]CS components isolated from HA-PGs after heparinase treatment showed that a major portion of these contained one CS chain only. Calculations of the number of CS and heparin chains in AT-binding PGs, based on polysaccharide composition and polysaccharide chain length, indicate that all heparin-containing PGs are hybrids.

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