NUCLEIC ACID SEQUENCES AND EXPRESSION SYSTEMS FOR HEPARINASEII AND HEPARINASE III DERIVED FROM FLAVOBACTERIUM HEPARINUM

摘要

The present invention describes the isolation and sequence of genes from Flavobacterium heparinum encoding heparin and heparan sulfate degrading ennzymes, heparinase II and heparinase III (EC 4.2.2.8). It further describes a method of expressing and an expression for heparinases I, II and III using a modified ribosome binding region derived from a promoter from glycosaminoglycan lyase genes of F. heparinum. Also, a multi-step protein purification method incorporating cell disruption, cation exchange chromatography, affinity chromatography and hydroxylapatite is outlined. Antibodies against a post-translational modification moiety common to Flavobacterium heparinum proteins and a method to obtain antibodies specific to these moieties and to the amino acid sequences of heparinases I,II and III are described.