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  4. GbpA as a secretion and affinity purification tag for an antimicrobial peptide produced in Vibrio natriegens
 
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2022
Journal Article
Title

GbpA as a secretion and affinity purification tag for an antimicrobial peptide produced in Vibrio natriegens

Abstract
Background: Vibrio natriegens is a Gram-negative bacterium that offers a greater metabolic capacity than Escherichia coli for the production of recombinant proteins. This potential includes a low minimum doubling time of 7 min and a high maximum glucose uptake rate of 3.9 g*g−1*h−1. We therefore tested the ability of V. natriegens to produce the insect metalloprotease inhibitor (IMPI), an antimicrobial peptide, fused to the glucosamine-binding protein A (GbpA) secretion/purification tag, using the Vmax Express system. Results: The IMPI-GbpA fusion protein was secreted into the medium and could be purified directly from the fermentation supernatant by affinity chromatography, including on-column digestion with thrombin. We also modified the GbpA tag by deleting the second and third domains, which reduced the size of the tag while maintaining its functionality. This modification also increased the IMPI yield. Conclusions: The use of V. natriegens as an expression platform and GbpA for protein secretion and purification facilitates the inexpensive production of antimicrobial peptides. Our process achieved a higher volumetric yield than earlier attempts to produce recombinant IMPI in E. coli. However, the accumulation of IMPI causes V. natriegens growth arrest before the carbon source is depleted, suggesting it may be possible to achieve even greater productivity by further process optimization.
Author(s)
Schwarz, S.
Technische Hochschule Mittelhessen
Gerlach, Doreen
Fraunhofer-Institut für Molekularbiologie und Angewandte Oekologie IME  
Fan, R.
Technische Hochschule Mittelhessen
Czermak, P.
Technische Hochschule Mittelhessen
Journal
Electronic journal of biotechnology : EJB  
Open Access
DOI
10.1016/j.ejbt.2022.01.003
Additional full text version
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Language
English
Fraunhofer-Institut für Molekularbiologie und Angewandte Oekologie IME  
Keyword(s)
  • Affinity purification tag

  • Affinity tag

  • AMP

  • Antimicrobial peptide

  • GbpA

  • IMPI

  • Protein secretion

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