Regulation and expression of human Fabs under the control of the Escherichia coli arabinose promoter, PBAD.

BACKGROUND

The L-arabinose operon from E. coli contains an inducible promoter PBAD which has been extensively studied for the control of gene expression. PBAD has a number of potential advantages over Plac, and has been used successfully to promote high level expression of recombinant proteins.

OBJECTIVES

The aim of this study was to investigate PBAD as an alternative system to Plac for the bacterial expression of recombinant Fabs.

STUDY DESIGN

The promoter PBAD from the E. coli arabinose operon araBAD and the gene encoding the regulator of this promoter, were cloned into the phagemid expression vector MCO1. Expression of human recombinant tetanus toxoid (TT) and c-erbB2 Fabs under the control of PBAD was compared at two induction temperatures with the same Fabs produced under the control of Plac.

RESULTS

Expression of TT and c-erbB2 Fabs under the control of PBAD was comparable to Fab expression from Plac. However, highly expressed TT Fab under the control of PBAD was localised to the soluble periplasmic fraction whereas under the control of Plac, there was greater leakage of Fab into the culture supernatant. In addition, Fab expression from PBAD could be more tightly repressed than from Plac.

CONCLUSION

PBAD is a useful and cheaply inducible alternative to the more commonly used Plac for the rapid expression of soluble recombinant human antibody fragments.