Abstract
Glycosaminoglycan (GAG) lyase is an effective tool for the structural and functional studies of glycosaminoglycans and preparation of functional oligosaccharides. A new GAG lyase from Microbacterium sp. H14 was cloned, expressed, purified, and characterized, with a molecular weight of approximately 85.9 kDa. The deduced lyase HCLaseM belonged to the polysaccharide lyase (PL) family 8. Based on the phylogenetic tree, HCLaseM could not be classified into the existing three subfamilies of this family. HCLaseM showed almost the same enzyme activity towards hyaluronan (HA), chondroitin sulfate A (CS-A), CS-B, CS-C, and CS-D, which was different from reported GAG lyases. HCLaseM exhibited the highest activities to both HA and CS-A at its optimal temperature (35 °C) and pH (pH 7.0). HCLaseM was stable in the range of pH 5.0–8.0 and temperature below 30 °C. The enzyme activity was independent of divalent metal ions and was not obviously affected by most metal ions. HCLaseM is an endo-type enzyme yielding unsaturated disaccharides as the end products. The facilitated diffusion effect of HCLaseM is dose-dependent in animal experiments. These properties make it a candidate for further basic research and application.
Highlights
Hyaluronan (HA) and chondroitin sulfate/dermatan sulfate (CS/DS) belong to the class of negatively charged linear heteropolysaccharides named glycosaminoglycans (GAGs) [1,2], which are widely distributed on the surface of cells and connective tissue of animals
According the Carbohydrate-Active enZYmes (CAZy) database, glycosaminoglycan lyases are classified into the enzymes of polysaccharide lyase (PL)-8, PL-16, PL-29 and other families
The enzymes of the PL-8 family are an important component of the GAG lyases, which can be divided into three subfamilies by phylogenetic analysis
Summary
Hyaluronan (HA) and chondroitin sulfate/dermatan sulfate (CS/DS) belong to the class of negatively charged linear heteropolysaccharides named glycosaminoglycans (GAGs) [1,2], which are widely distributed on the surface of cells and connective tissue of animals. Enzymes degrading GAGs are associated with the regulation of various cellular processes, such as proliferation, differentiation, migration, and adhesion [3,4]. GAG-based therapeutic agents have important application prospects. According the Carbohydrate-Active enZYmes (CAZy) database, glycosaminoglycan lyases are classified into the enzymes of PL-8, PL-16, PL-29 and other families. The enzymes of the PL-8 family can degrade more types of substrates. The enzymes of the PL-8 family are an important component of the GAG lyases, which can be divided into three subfamilies by phylogenetic analysis
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