Production in Escherichia coli of porcine type-I collagenase as a fusion protein with beta-galactosidase

Abstract

Porcine type-I collagenase (Colg-1) was produced as a fusion protein in Escherichia coli using the pAX5 expression vector. The fusion protein consists of beta-galactosidase at the N terminus joined to a collagen hinge region and a blood-coagulation factor Xa cleavage site linked to Colg-1. Recombinant collagenase (reColg-1) was biologically active in the form of a fusion protein and could be released by treatment with factor Xa to yield Colg-1 with the authentic N terminus (phenylalanine) found in vivo. The results show that reColg-1 produced in E. coli is folded correctly, cleaves type-I collagen into 1/4 and 3/4 fragments at the characteristic Colg-sensitive site, and is produced at high enough levels to generate a source of recombinant enzyme for x-ray crystallography studies.