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Translational level is a critical factor for the secretion of heterologous proteins in Escherichia coli

Nature Biotechnology volume 14, pages 629–634 (1996)Cite this article

Abstract

A method for enhancing the secretion of heterologous proteins in Escherichia coli by optimizing, as opposed to simply maximizing, the translational level of a given protein is described. Random alteration of the translational initiation region (TIR) of the Heat-Stable Enterotoxin II (STII) signal sequence resulted in a library of vectors with varied translational strengths. Subsequent screening of this library using E. coli alkaline phosphatase as a reporter led to the selection of several TIR variants covering a 10-fold range of translational strength. These TIR variants, in combination with several previously generated variants, are shown to dramatically improve the secretion of a number of heterologous proteins. In fact, the heterologous proteins tested required a narrow translational range for optimal high-level secretion into the periplasm. Interestingly, the secretion of two native E. coli proteins was unaffected by TIR strength when tested over an identical range. The dependence of secretion on a narrow translational level demonstrates its critical role in the secretion of heterologous proteins in E. coli.

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Author notes

  1. Laura C. Simmons: e-mail: simmons@gene.com.

Affiliations

  1. Department of Molecular Biology, Genentech, Inc., 460 Point San Bruno Blvd., South San Francisco, CA, 94080

    Laura C. Simmons & Daniel G. Yansura

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  1. Laura C. Simmons
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  2. Daniel G. Yansura
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Simmons, L., Yansura, D. Translational level is a critical factor for the secretion of heterologous proteins in Escherichia coli. Nat Biotechnol 14, 629–634 (1996). https://doi.org/10.1038/nbt0596-629

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  • DOI: https://doi.org/10.1038/nbt0596-629

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