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. 1990 Jan;87(1):142-6.
doi: 10.1073/pnas.87.1.142.

Expression of a cloned streptavidin gene in Escherichia coli

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Expression of a cloned streptavidin gene in Escherichia coli

T Sano et al. Proc Natl Acad Sci U S A. 1990 Jan.

Abstract

We describe the construction of systems for expressing the cloned streptavidin gene in Escherichia coli. Although the streptavidin gene is extremely lethal to the host cells, because of the strong biotin binding of the gene product, the gene was expressed efficiently in E. coli by using T7 RNA polymerase/T7 promoter expression systems. The expressed streptavidin accumulated to more than 35% of the total cell protein. The expressed streptavidin was insoluble in the cell. However, after solubilization by dialysis against 6 M guanidine hydrochloride (pH 1.5) and removal of guanidine hydrochloride by dialysis, the protein became soluble and renatured. This simple procedure yielded streptavidin purified almost to homogeneity. The purified streptavidin bound 3.5-3.9 molecules of biotin per molecule, indicating that it had almost full biotin-binding ability. Some of the purified streptavidin molecules aggregated into oligomers, suggesting that the C-terminal region of the molecule, present in our material but absent in typical preparations, may be responsible for the aggregation.

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