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Extracellular accumulation of recombinant proteins fused to the carrier protein YebF in Escherichia coli

Nature Biotechnology volume 24, pages 100–104 (2006)Cite this article

Abstract

Bacterial protein secretion is important in the life cycles of most bacteria, in which it contributes to the formation of pili and flagella and makes available extracellular enzymes to digest polymers for nutritional purposes and toxins to kill host cells in infections of humans, animals and plants. It is generally accepted that nonpathogenic laboratory strains of Escherichia coli, particularly K12 strains, do not secrete proteins into the extracellular medium under routine growth conditions1,2. In this study, we report that commonly used laboratory strains secrete YebF, a small (10.8 kDa in the native form), soluble endogenous protein into the medium, challenging the status quo view that laboratory strains do not secrete proteins to the medium. We further show that 'passenger' proteins linked to the carboxyl end of YebF are efficiently secreted. The function of YebF is unknown, but its use as a carrier for transgenic proteins provides a tool to circumvent toxicity and other contamination issues associated with protein production in E. coli.

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Figure 1: Subcellular localization and secretion of E. coli YebF.
Figure 2: Immunoblotting of YebF and CRP in HB101 (pYebFH6/MS) cells and the medium.
Figure 3: Examination of outer membrane leakage in E. coli cells expressing YebF protein.
Figure 4: YebF as the carrier protein to produce hIL2 or α-amylase in the medium.

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Acknowledgements

J.H.W. is a Canada Research Chair in Membrane Biochemistry. This work was supported by the Canadian Institutes of Health Research and the Alberta Heritage Foundation for Medical Research. We thank the following people for their valuable contribution to this study. Len Wiebe and Aihua Zhou provided technical assistance with the hIL-2 bioassay. Chris Bleackley, Irene Shostak and Jonathan Hooton provided the materials and methods for the hIL-2 bioassay. Lorne Burke and Paul Semchuk provided technical support for HPLC and mass spectrometry. We thank Biomira for kindly allowing us to use plasmid pBM806.

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  1. Department of Biochemistry, Membrane Protein Research Group, 474 Medical Sciences Building, University of Alberta, Edmonton, T6G 2H7, Alberta, Canada

    Guijin Zhang, Stephen Brokx & Joel H Weiner

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  1. Guijin Zhang
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Correspondence to Joel H Weiner.

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Zhang, G., Brokx, S. & Weiner, J. Extracellular accumulation of recombinant proteins fused to the carrier protein YebF in Escherichia coli. Nat Biotechnol 24, 100–104 (2006). https://doi.org/10.1038/nbt1174

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