Chondroitinase ABC I (ChSase ABC I) is a kind of lyases that can degrade glycosaminoglycans such as chondroitin sulfate, chondroitin and hyaluronic acid into oligosaccharides and unsaturated disaccharides. The ChSase ABC I gene was linked to maltose binging protein (MBP) gene using two linker peptides including FFFFF and RRRRR, respectively, cloned into pMAL-c2X vector, and successfully expressed in Escherichia coli BL21 (DE3). The results showed that the enzymatic activity and specific enzyme activity of the recombinase MBP-FFFFF-ChSase ABC I were 716.5 IU/L fermentation broth and 1.15 IU/mg protein, respectively, and the recombinase MBP-RRRRR-ChSase ABC I were 741.0 IU/L fermentation broth and 1.13 IU/mg protein, respectively. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) results showed that the molecular weights of the recombinases MBP-FFFFF-ChSase ABC I and MBP-RRRRR-ChSase ABC I were 130 kDa, and they were soluble proteins.