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Production of halophilic proteins using Haloferax volcanii H1895 in a stirred-tank bioreactor

Strillinger, Eva; Gr�tzinger, Stefan Wolfgang; Allers, Thorsten; Eppinger, J�rg; Weuster-Botz, Dirk

Authors

Eva Strillinger

Stefan Wolfgang Gr�tzinger

THORSTEN ALLERS THORSTEN.ALLERS@NOTTINGHAM.AC.UK
Professor of Archaeal Genetics

J�rg Eppinger

Dirk Weuster-Botz



Abstract

The success of biotechnological processes is based on the availability of efficient and highly specific biocatalysts, which can satisfy industrial demands. Extreme and remote environments like the deep brine pools of the Red Sea represent highly interesting habitats for the discovery of novel halophilic and thermophilic enzymes. Haloferax volcanii constitutes a suitable expression system for halophilic enzymes obtained from such brine pools. We developed a batch process for the cultivation of H. volcanii H1895 in controlled stirred-tank bioreactors utilising knockouts of components of the flagella assembly system. The standard medium Hv-YPC was supplemented to reach a higher cell density. Without protein expression, cell dry weight reaches 10 g L?1. Two halophilic alcohol dehydrogenases were expressed under the control of the tryptophanase promoter p.tna with 16.8 and 3.2 mg gCDW ?1, respectively, at a maximum cell dry weight of 6.5 g L?1. Protein expression was induced by the addition of l-tryptophan. Investigation of various expression strategies leads to an optimised two-step induction protocol introducing 6 mM l-tryptophan at an OD650 of 0.4 followed by incubation for 16 h and a second induction step with 3 mM l-tryptophan followed by a final incubation time of 4 h. Compared with the uncontrolled shaker-flask cultivations used until date, dry cell mass concentrations were improved by a factor of more than 5 and cell-specific enzyme activities showed an up to 28-fold increased yield of the heterologous proteins.

Citation

Strillinger, E., Grötzinger, S. W., Allers, T., Eppinger, J., & Weuster-Botz, D. (2016). Production of halophilic proteins using Haloferax volcanii H1895 in a stirred-tank bioreactor. Applied Microbiology and Biotechnology, 100(3), 1183-1195. https://doi.org/10.1007/s00253-015-7007-1

Journal Article Type Article
Acceptance Date Sep 12, 2015
Online Publication Date Oct 1, 2015
Publication Date 2016-02
Deposit Date Feb 7, 2019
Journal Applied Microbiology and Biotechnology
Print ISSN 0175-7598
Electronic ISSN 1432-0614
Publisher BMC
Peer Reviewed Peer Reviewed
Volume 100
Issue 3
Pages 1183-1195
DOI https://doi.org/10.1007/s00253-015-7007-1
Public URL https://nottingham-repository.worktribe.com/output/1523521
Publisher URL https://link.springer.com/article/10.1007%2Fs00253-015-7007-1