Expression of a β-Lactamase Preproinsulin Fusion Protein in Escherichia Coli

The level of expression of a β-lactamase human preproinsulin fusion protein 1 in Escherichia coli was increased by plasmid and host manipulations. The initial expression level (0.01% of total protein) was increased two- to three-fold by replacing the p-lactamase promoter with two strong promoters: t...

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Bibliographic Details
Published in:Bio/Technology 1984-02, Vol.2 (2), p.165-168
Main Authors: Emerick, Anne W., Bertolani, Barbara L., Ben-Bassat, A., White, Thomas J., Konrad, Michael W.
Format: Article
Language:English
Subjects:
Agriculture
Bioinformatics
Biomedical and Life Sciences
Biomedical Engineering/Biotechnology
Biomedicine
Biotechnology
Life Sciences
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Summary:The level of expression of a β-lactamase human preproinsulin fusion protein 1 in Escherichia coli was increased by plasmid and host manipulations. The initial expression level (0.01% of total protein) was increased two- to three-fold by replacing the p-lactamase promoter with two strong promoters: trp from Serratia marcescens and lac UV5 from E. coli. Replacement of the β-lactamase ribosome binding site (RBS) with a trp RBS enhanced the yield to a total of 0.16% of the total cell protein. Proinsulin was unstable, with a half-life of 14 minutes at 37°C. Proinsulin expression was increased three-fold (to about 0.5% of the total cell protein) by incubating the host at 30°C rather than 37°C.
ISSN:0733-222X
1087-0156
1546-1696
DOI:10.1038/nbt0284-165