Biotechnol. Appl. Biochem., 38, 87-93 (2004)

Evaluation of inducible promoters on the secretion of a ZZ-proinsulin fusion protein in Escherichia coli

Mergulhão, F. J. M., Monteiro, G. A., Larsson, G., Bostrom, M., Farewell, A., Nystrom, T., Cabral, J. M. S., Taipa, M. A.

Abstract

Four inducible promoters, uspA, uspS, lacUV5 and malK were evaluated in the expression of the fusion protein ZZ-Proinsulin by Escherichia coli to select the most appropriate expression system (promoter and culture medium) for secretion to the periplasmic space and culture medium. All the expression vectors contain the RNaselll cleavage site to ensure that mRNA translation rate is independent of 5' untranslated regions thus making promoter strength comparisons more accurate. The highest ZZ- Proinsulin secretion yields reached 6.2 mg/g dry cell weight in the periplasmic space and 2.6 mg/g dry cell weight in the culture medium using the malK promoter. It was also demonstrated that the use of M9 minimal medium favours secretion.

Keywords: E. coli; induction; malK; stress; recombinant systems

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