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

Handle URI:
http://hdl.handle.net/10754/579494
Title:
Production of halophilic proteins using Haloferax volcanii H1895 in a stirred-tank bioreactor
Authors:
Strillinger, Eva ( 0000-0003-1159-6981 ) ; Grötzinger, Stefan W.; Allers, Thorsten; Eppinger, Jorg ( 0000-0001-7886-7059 ) ; Weuster-Botz, Dirk
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.
KAUST Department:
Physical Sciences and Engineering (PSE) Division; Biological & Organometallic Catalysis Laboratory; KAUST Catalysis Center (KCC); Computer, Electrical and Mathematical Sciences and Engineering (CEMSE) Division; Computational Bioscience Research Center (CBRC)
Citation:
Production of halophilic proteins using Haloferax volcanii H1895 in a stirred-tank bioreactor 2015 Applied Microbiology and Biotechnology
Publisher:
Springer Science + Business Media
Journal:
Applied Microbiology and Biotechnology
Issue Date:
1-Oct-2015
DOI:
10.1007/s00253-015-7007-1
Type:
Article
ISSN:
0175-7598; 1432-0614
Additional Links:
http://link.springer.com/10.1007/s00253-015-7007-1
Appears in Collections:
Articles; Physical Sciences and Engineering (PSE) Division; KAUST Catalysis Center (KCC); Computational Bioscience Research Center (CBRC); Computer, Electrical and Mathematical Sciences and Engineering (CEMSE) Division

Full metadata record

DC FieldValue Language
dc.contributor.authorStrillinger, Evaen
dc.contributor.authorGrötzinger, Stefan W.en
dc.contributor.authorAllers, Thorstenen
dc.contributor.authorEppinger, Jorgen
dc.contributor.authorWeuster-Botz, Dirken
dc.date.accessioned2015-10-08T11:27:50Zen
dc.date.available2015-10-08T11:27:50Zen
dc.date.issued2015-10-01en
dc.identifier.citationProduction of halophilic proteins using Haloferax volcanii H1895 in a stirred-tank bioreactor 2015 Applied Microbiology and Biotechnologyen
dc.identifier.issn0175-7598en
dc.identifier.issn1432-0614en
dc.identifier.doi10.1007/s00253-015-7007-1en
dc.identifier.urihttp://hdl.handle.net/10754/579494en
dc.description.abstractThe 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.en
dc.language.isoenen
dc.publisherSpringer Science + Business Mediaen
dc.relation.urlhttp://link.springer.com/10.1007/s00253-015-7007-1en
dc.rightsThe final publication is available at Springer via http://dx.doi.org/10.1007/s00253-015-7007-1en
dc.subjectGene expressionen
dc.subjectHalophileen
dc.subjectHaloferax volcaniien
dc.subjectStirred-tank bioreactoren
dc.subjectBiocatalysisen
dc.subjectBiotechnologyen
dc.titleProduction of halophilic proteins using Haloferax volcanii H1895 in a stirred-tank bioreactoren
dc.typeArticleen
dc.contributor.departmentPhysical Sciences and Engineering (PSE) Divisionen
dc.contributor.departmentBiological & Organometallic Catalysis Laboratoryen
dc.contributor.departmentKAUST Catalysis Center (KCC)en
dc.contributor.departmentComputer, Electrical and Mathematical Sciences and Engineering (CEMSE) Divisionen
dc.contributor.departmentComputational Bioscience Research Center (CBRC)en
dc.identifier.journalApplied Microbiology and Biotechnologyen
dc.eprint.versionPost-printen
dc.contributor.institutionInstitute of Biochemical Engineering, Department of Mechanical Engineering, Technische Universität München, Munich, Germanyen
dc.contributor.institutionInstitute of Genetics, School of Biology, University of Nottingham, Queen’s Medical Centre, Nottingham, UKen
dc.contributor.affiliationKing Abdullah University of Science and Technology (KAUST)en
kaust.authorEppinger, Jorgen
kaust.authorGrötzinger, Stefan W.en
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