Expression of the heparinase gene from Flavobacterium heparinum in Escherichia coli and its enzymatic properties

Heparinase has an important application in the preparation of low-molecular-weight heparins by heparin enzymolysis. A heparinase gene from Flavobacterium heparinum was cloned and expressed in Escherichia coli BL21 in order to enhance its activity. The expressed heparinase was purified to homogeneity by a metal chelating affinity column and its enzymatic properties were evaluated. A maximal heparinase activity of 1061 IU/L toward the substrate heparin was achieved when the recombinant strain was induced with 0.5 mM isopropyl-β-D-thiogalactoside at 28 °C for 9 h. The optimal temperature and pH of heparinase were 30 °C and 7.0, respectively. The recombinant heparinase was heat-unstable and had a higher stability at pHs from 7.0 to 10.0. Observed activities of heparinase were the highest in the presence of Ca(2+) and Cu(2+) and the lowest in the presence of Mn(2+) and Pb(2+). These results lay a good foundation for the preparation of LMWHs by heparin enzymolysis.