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Dibenzothiophene biodesulfurization pathway improvement using diagnostic GFP fusions
David S Reichmuth1, Harvey W Blanch, Jay D Keasling
1Department of Chemical Engineering, University of California, Berkeley, CA 94720, USA.
Biotechnology and Bioengineering
|September 25, 2004
Summary
Researchers enhanced the biodesulfurization pathway by increasing DszB enzyme production. This resulted in a ninefold increase in the rate of removing sulfur from fossil fuels, improving fuel processing.
Area of Science:
- Biotechnology
- Environmental Science
- Microbial Engineering
Background:
- Dibenzothiophene biodesulfurization offers a promising method for processing sulfur-containing fossil fuels.
- The efficiency of this pathway is often limited by low levels of the DszB enzyme.
Purpose of the Study:
- To overcome the limitations of low DszB enzyme production in the dibenzothiophene biodesulfurization pathway.
- To enhance the overall rate of desulfurization in fossil fuels.
Main Methods:
- Mutagenesis of the 5' untranslated region of the dszB gene using degenerate oligonucleotides.
- Utilized translational fusions with green fluorescent protein (GFP) to screen for increased dszB translation.
- Screened 96 mutants to identify those with enhanced DszB activity.
Main Results:
- Several mutants exhibited increased green fluorescence, indicating higher dszB translation.
- Two mutants demonstrated significantly increased DszB enzyme activity.
- Cotransformation with the full dszABC operon and mutant dszB resulted in a ninefold increase in desulfurization rate compared to native dszB.
Conclusions:
- Targeted mutations in the dszB 5' untranslated region effectively enhance DszB production and activity.
- This strategy significantly improves the efficiency of the dibenzothiophene biodesulfurization pathway.
- The enhanced pathway holds potential for more effective processing of sulfur-containing fossil fuels.