A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus

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 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 beta-glucosidase gene revealed that the pilA4 promoter is highly active under optimal growth conditions at 68 degrees C and downregulated during growth at 80 degrees 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 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.

Automated summary

In this study, the pilA4 promoter of Thermus thermophilus was characterized and found to be highly active under optimal growth conditions and downregulated at the highest growth temperature. The promoter activity was significantly increased in minimal medium compared to complex medium. Furthermore, the suitability of the pilA4 promoter for heterologous production of a thermostable enzyme was demonstrated.