N-ACETYL-HEPAROSAN LYASE OF ESCHERICHIA-COLI K5 - GENE CLONING AND EXPRESSION

The structure of the capsular polysaccharide of Escherichia coli K5 is identical to that of N-acetyl-heparosan, a nonsulfated precursor of h eparin, which makes this E. coli antigen an attractive starting point for the chemical synthesis of analogs of low-molecular-weight heparin. This polysaccharide is synthesized as a high-molecular-weight molecul e that can be depolymerized by an enzyme displaying endo-beta-eliminas e activity. The eliminase-encoding gene, designated elmA, has been clo ned from E. coli K5 by expression in E. coli K-12. The K-12 genome is devoid of the elmA sequence. The elmA gene product is 820 amino acids long. Active recombinant eliminase is produced by K-12 cells in both c ell-bound and secreted forms. Deletion analyses have shown that the C terminus and the N terminus are required for activity and secretion, r espectively.