The substrate specificity of heparan sulphate lyase and heparin lyase from Flavobacterium heparinum

Abstract

The substrate specificity of heparan sulphate lyase and heparin lyase has been examined by using various oligosaccharides produced by deaminative cleavage or periodate oxidation—base-catalysed elimination of heparan sulphate. The saccharides were separated by gel and ion-exchange chromatography into fractions having high proportions of the following linkages: 2-acetamido-2-deoxy- d-glucosyl→ d-glucuronic acid (type I), 2-deoxy-2-sulphoamino- d-glucosyl→ d-glucuronic acid (type II), 2-deoxy-2-sulphoamino- d-glucosyl→ l-iduronic acid (type III), or 2-deoxy-2-sulphoamino- d-glucosyl→2- O-sulpho- l-iduronic acid (type IV). Heparan sulphate lyase cleaved heparan sulphate preparations containing high proportions of type I linkages. In contrast, oligosaccharides obtained after deaminative cleavage and rich in the type I linkage were poor substrates; longer saccharides were better substrates than shorter ones. Saccharides generated via periodate oxidation were cleaved when the type I linkage was present but resistant when the linkages were of types II-IV. The presence of ester sulphate on the 2-acetamido-2-deoxy- d-glucose residue in a type I linkage seems to hinder the enzyme. Heparan sulphate lyase was able to degrade intact chains to the same extent as did two cycles of periodate oxidation—base-catalysed elimination, suggesting that all the regular type I linkages were accessible to the enzyme. The heparin lyase could cleave saccharides which contained type IV linkages. When type II and III linkages were the only ones present, the saccharides were resistant. This enzyme also appears to require ester-sulphation of the 2-deoxy-2-sulphoamino- d-glucose residue.