Abstract
BackgroundPreviously we have developed a butanol tolerant mutant of Clostridium acetobutylicum Rh8, from the wild type strain DSM 1731. Strain Rh8 can tolerate up to 19 g/L butanol, with solvent titer improved accordingly, thus exhibiting industrial application potential. To test if strain Rh8 can be used for production of high level mixed alcohols, a single secondary alcohol dehydrogenase from Clostridium beijerinckii NRRL B593 was overexpressed in strain Rh8 under the control of thl promoter.ResultsThe heterogenous gene sADH was functionally expressed in C. acetobutylicum Rh8. This simple, one-step engineering approach switched the traditional ABE (acetone-butanol-ethanol) fermentation to IBE (isopropanol-butanol-ethanol) fermentation. The total alcohol titer reached 23.88 g/l (7.6 g/l isopropanol, 15 g/l butanol, and 1.28 g/l ethanol) with a yield to glucose of 31.42%. The acid (butyrate and acetate) assimilation rate in isopropanol producing strain Rh8(psADH) was increased.ConclusionsThe improved butanol tolerance and the enhanced solvent biosynthesis machinery in strain Rh8 is beneficial for production of high concentration of mixed alcohols. Strain Rh8 can thus be considered as a good host for further engineering of solvent/alcohol production.
Highlights
We have developed a butanol tolerant mutant of Clostridium acetobutylicum Rh8, from the wild type strain DSM 1731
Functional expression of secondary alcohol dehydrogenase in C. acetobutylicum Rh8 To test the genetic manipulation feasibility of strain Rh8, both methylated and unmethylated shuttle vector pIMP1 were transformed into this mutant strain
The results showed that strain Rh8 could accept the methylated plasmid with normal transformation efficiency (104 transformants per μg DNA)
Summary
We have developed a butanol tolerant mutant of Clostridium acetobutylicum Rh8, from the wild type strain DSM 1731. Titer, and productivity of ABE fermentation, various engineering strategies have been developed These includes engineering central carbon flux redistribution [2,3,4,5,6], engineering regulatory mechanism [7,8], engineering phenotypic properties [9], and engineering butanol tolerance [10,11]. A butanol tolerant C. acetobutylicum mutant Rh8 was obtained by chemical mutagenesis and genome shuffling This mutant showed a 46% improved butanol and 20% improved solvent titer, compared to that of the wild type strain DSM 1731 [15]. These comprehensive characterizations suggest that strain Rh8 might be considered as an interesting host for further improvement of solvent productivity
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