A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus.
Gene expression
Promoters
Thermophiles
Thermozymes
Thermus thermophilus HB27
Journal
Microbial cell factories
ISSN: 1475-2859
Titre abrégé: Microb Cell Fact
Pays: England
ID NLM: 101139812
Informations de publication
Date de publication:
19 Sep 2023
19 Sep 2023
Historique:
received:
27
07
2023
accepted:
02
09
2023
medline:
21
9
2023
pubmed:
20
9
2023
entrez:
19
9
2023
Statut:
epublish
Résumé
Enzymes from thermophiles are of great interest for research and bioengineering due to their stability and efficiency. Thermophilic expression hosts such as Thermus thermophilus [T. thermophilus] can overcome specific challenges experienced with protein production in mesophilic expression hosts, such as leading to better folding, increased protein stability, solubility, and enzymatic activity. However, available inducible promoters for efficient protein production in T. thermophilus HB27 are limited. In this study, we characterized the pilA4 promoter region and evaluated its potential as a tool for production of thermostable enzymes in T. thermophilus HB27. Reporter gene analysis using a promoterless β-glucosidase gene revealed that the pilA4 promoter is highly active under optimal growth conditions at 68 °C and downregulated during growth at 80 °C. Furthermore, growth in minimal medium led to significantly increased promoter activity in comparison to growth in complex medium. Finally, we proved the suitability of the pilA4 promoter for heterologous production of thermostable enzymes in T. thermophilus by producing a fully active soluble mannitol-1-phosphate dehydrogenase from Thermoanaerobacter kivui [T. kivui], which is used in degradation of brown algae that are rich in mannitol. Our results show that the pilA4 promoter is an efficient tool for gene expression in T. thermophilus with a high potential for use in biotechnology and synthetic biology applications.
Sections du résumé
BACKGROUND
BACKGROUND
Enzymes from thermophiles are of great interest for research and bioengineering due to their stability and efficiency. Thermophilic expression hosts such as Thermus thermophilus [T. thermophilus] can overcome specific challenges experienced with protein production in mesophilic expression hosts, such as leading to better folding, increased protein stability, solubility, and enzymatic activity. However, available inducible promoters for efficient protein production in T. thermophilus HB27 are limited.
RESULTS
RESULTS
In this study, we characterized the pilA4 promoter region and evaluated its potential as a tool for production of thermostable enzymes in T. thermophilus HB27. Reporter gene analysis using a promoterless β-glucosidase gene revealed that the pilA4 promoter is highly active under optimal growth conditions at 68 °C and downregulated during growth at 80 °C. Furthermore, growth in minimal medium led to significantly increased promoter activity in comparison to growth in complex medium. Finally, we proved the suitability of the pilA4 promoter for heterologous production of thermostable enzymes in T. thermophilus by producing a fully active soluble mannitol-1-phosphate dehydrogenase from Thermoanaerobacter kivui [T. kivui], which is used in degradation of brown algae that are rich in mannitol.
CONCLUSIONS
CONCLUSIONS
Our results show that the pilA4 promoter is an efficient tool for gene expression in T. thermophilus with a high potential for use in biotechnology and synthetic biology applications.
Identifiants
pubmed: 37726752
doi: 10.1186/s12934-023-02192-1
pii: 10.1186/s12934-023-02192-1
pmc: PMC10507856
doi:
Substances chimiques
Fimbriae Proteins
147680-16-8
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
187Subventions
Organisme : Deutsche Forschungsgemeinschaft
ID : AV 9/6-2
Organisme : Bundesministerium für Bildung und Forschung
ID : ThermoSynCon
Informations de copyright
© 2023. BioMed Central Ltd., part of Springer Nature.
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