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

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