Introductions
Heparinases are a class of polysaccharide cleaving enzymes that act on heparin or acetyl heparin molecules. Heparinases include both microbially produced enzymes that degrade heparin or acetyl heparan sulfate by cleavage and endogenous (heparanase-like) enzymes produced in animals that degrade acetyl heparan sulfate by hydrolysis.
Source
Heparanase has been isolated mainly from some bacteria that use heparin as a carbon source, and was originally derived from Flavobacterium heparinum. In addition, heparinase has been found in many microorganisms such as Corynebacterium sp. However, Flavobacterium heparinum is still the only source of commercial heparinase. Three heparinases have been isolated and purified from Heparinibacterium heparinum, heparinase I, II and III, which have different enzymatic properties. Among the three heparinases purified from Flavobacterium heparinum, only the structure of heparinase II has been resolved and it has the least substrate specificity; heparinase I is most suitable for the preparation of low molecular weight heparin.
Properties
Heparanase selectively shears the α(1-4) glycosidic bond between glucosamine and glyoxylate in sulfated heparin glycans, and three different heparanases (enzyme I, enzyme II, and enzyme III) have very different specificities. Heparanase I breaks the linkage between heparin and acetyl heparan sulfate (relative activity about 3:1) in glucosamine and o-aidulose sulfate, and the product is mainly disaccharide. This enzyme also cuts the antithrombin III binding pentasaccharide site in the heparin molecule.
Activity
Heparanase selectively shears the α(1-4) glycosidic bond between glucosamine and glucuronide in sulfated heparan glycans, and three different heparanases (enzyme I, enzyme II, and enzyme III) have very different specificities. Heparanase III shears heparan sulfate at the 1-4 linkage site between hexosamine and glucuronide, producing mainly disaccharides. This enzyme does not act on heparin and low molecular weight heparin.
Applications
Heparinase has many important pharmaceutical applications, the main uses are as follows.
(1) Preparation of low-molecular and ultra-low-molecular heparin
Low- and ultra-low-molecular-weight heparins have anticoagulant properties and special effects in inhibiting vascular proliferation, preventing tumor metastasis, treating cancer, and anti-allergy. The production of low molecular weight heparin by heparinase is an ideal industrial production method because it is mild, environmentally friendly and does not act on the sulfate group.
(2) Elimination of heparin from circulating blood in vitro
Some critically ill patients, such as uremic patients, require extracorporeal circulation of blood during surgery, during which heparin is added to prevent blood clotting. To prevent heparin from remaining in the blood and affecting blood clotting, fisetin is added clinically to neutralize heparin. Clinical studies have found that fisetin severely damages platelets and affects the stability of arterial blood pressure, whereas heparinase has no such side effects and is therefore an ideal alternative material. It should be noted that since heparinase also degrades the extracellular matrix, the heparinase used for heparinization removal of blood also needs to be immobilized, and it is required that the heparinase must not be immunogenic.
(3) Determination of the precise structure of heparin
The heparin to be analyzed is first degraded by purified heparinase from Heparin Flavobacterium to obtain the tetrasaccharide product, and then deep degradation is carried out by purified acetyl heparinase, also from Heparin Flavobacterium, and the structure of the corresponding heparin can be inferred by analysis of the final product.
