Disulfide bond formation and folding of plant peroxidases expressed as inclusion body protein in Escherichia coli thioredoxin reductase negative strains

Escherichia coli is widely used for the production of proteins, which are o f interest in structure and function studies. The folding yield of inclusio n body protein is, however, generally low (a few percent) for proteins such as the plant and fungal peroxidases, which contain four disulfide bonds, t wo Ca2+ ions, and a heme group. We have studied the expression yield and fo lding efficiency of (i) a novel Arabidopsis thaliana peroxidase, ATP N; and (ii) barley grain peroxidase, BP 1. The expression yield ranges from 0 to 60 mu g/ml of cell culture depending on the peroxidase gene and the vector/ host combination. The choice off. coli strain in particular affects the yie ld of active peroxidase obtained in the folding step. Thus, the yield of ac tive ATP N peroxidase can be increased 50-fold by using thioredoxin reducta se negative strains, which facilitate the formation of disulfide bonds in i nclusion body protein. (C) 1999 Academic Press.