Use of the human hepcidin gene to build a positive-selection vector for periplasmic expression in Escherichia coli

Jérome Haustant; Annesha Sil; Christopher Maillo-Rius; Agnès Hocquellet; Patricia Costaglioli; Bertrand Garbay; Wilfrid Dieryck

doi:10.1016/j.ab.2016.02.002

Use of the human hepcidin gene to build a positive-selection vector for periplasmic expression in Escherichia coli

Jérome Haustant, Annesha Sil, Christopher Maillo-Rius, Agnès Hocquellet, Patricia Costaglioli, Bertrand Garbay, Wilfrid Dieryck (Corresponding Author)

Research output: Contribution to journal › Article › peer-review

2 Citations (Scopus)

Abstract

Recombinant proteins are often produced in the periplasm of Escherichia coli because this facilitates the purification process. The oxidizing environment favors the formation of disulfide bridges. We showed that the periplasmic expression of the human hormone hepcidin 25 (Hep25) fused to the maltose-binding protein (MBP) resulted in cell death. This toxicity was not observed when MBP–Hep25 accumulated in the bacterial cytoplasm, or when Hep25 was addressed to the periplasm without the MBP tag. We then modified the periplasmic expression vector pMALp2E to create pMALp2EH, a positive-selection vector with Hep25 as counterselection gene.

Original language	English
Pages (from-to)	35-37
Number of pages	3
Journal	Analytical Biochemistry
Volume	500
Early online date	10 Feb 2016
DOIs	https://doi.org/10.1016/j.ab.2016.02.002
Publication status	Published - 1 May 2016
Externally published	Yes

Bibliographical note

Acknowledgments
This work was supported in part by grants from Bordeaux INP and Bordeaux University.

Keywords

positive selection
Hepcidin
expression vector
cloning

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Cite this

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title = "Use of the human hepcidin gene to build a positive-selection vector for periplasmic expression in Escherichia coli",

abstract = "Recombinant proteins are often produced in the periplasm of Escherichia coli because this facilitates the purification process. The oxidizing environment favors the formation of disulfide bridges. We showed that the periplasmic expression of the human hormone hepcidin 25 (Hep25) fused to the maltose-binding protein (MBP) resulted in cell death. This toxicity was not observed when MBP–Hep25 accumulated in the bacterial cytoplasm, or when Hep25 was addressed to the periplasm without the MBP tag. We then modified the periplasmic expression vector pMALp2E to create pMALp2EH, a positive-selection vector with Hep25 as counterselection gene.",

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doi = "10.1016/j.ab.2016.02.002",

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T1 - Use of the human hepcidin gene to build a positive-selection vector for periplasmic expression in Escherichia coli

AU - Haustant, Jérome

AU - Sil, Annesha

AU - Maillo-Rius, Christopher

AU - Hocquellet, Agnès

AU - Costaglioli, Patricia

AU - Garbay, Bertrand

AU - Dieryck, Wilfrid

N1 - Acknowledgments This work was supported in part by grants from Bordeaux INP and Bordeaux University.

PY - 2016/5/1

Y1 - 2016/5/1

N2 - Recombinant proteins are often produced in the periplasm of Escherichia coli because this facilitates the purification process. The oxidizing environment favors the formation of disulfide bridges. We showed that the periplasmic expression of the human hormone hepcidin 25 (Hep25) fused to the maltose-binding protein (MBP) resulted in cell death. This toxicity was not observed when MBP–Hep25 accumulated in the bacterial cytoplasm, or when Hep25 was addressed to the periplasm without the MBP tag. We then modified the periplasmic expression vector pMALp2E to create pMALp2EH, a positive-selection vector with Hep25 as counterselection gene.

AB - Recombinant proteins are often produced in the periplasm of Escherichia coli because this facilitates the purification process. The oxidizing environment favors the formation of disulfide bridges. We showed that the periplasmic expression of the human hormone hepcidin 25 (Hep25) fused to the maltose-binding protein (MBP) resulted in cell death. This toxicity was not observed when MBP–Hep25 accumulated in the bacterial cytoplasm, or when Hep25 was addressed to the periplasm without the MBP tag. We then modified the periplasmic expression vector pMALp2E to create pMALp2EH, a positive-selection vector with Hep25 as counterselection gene.

KW - positive selection

KW - Hepcidin

KW - expression vector

KW - cloning

U2 - 10.1016/j.ab.2016.02.002

DO - 10.1016/j.ab.2016.02.002

M3 - Article

SN - 0003-2697

VL - 500

SP - 35

EP - 37

JO - Analytical Biochemistry

JF - Analytical Biochemistry

ER -

Use of the human hepcidin gene to build a positive-selection vector for periplasmic expression in Escherichia coli

Abstract

Bibliographical note

Keywords

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