Acetone production with metabolically engineered strains of Acetobacterium woodii

• Published in: Metabolic engineering 2016-07, Vol.36, p.37-47
• Main Authors: Hoffmeister, Sabrina, Gerdom, Marzena, Bengelsdorf, Frank R., Linder, Sonja, Flüchter, Sebastian, Öztürk, Hatice, Blümke, Wilfried, May, Antje, Fischer, Ralf-Jörg, Bahl, Hubert, Dürre, Peter
• Format: Article
• Language: English
• Subjects: Acetates - metabolism; Acetobacterium - physiology; Acetobacterium woodii; Acetone; Acetone - isolation & purification; Acetone - metabolism; Biosynthetic Pathways - physiology; Carbon dioxide; Carbon Dioxide - metabolism; CoA transferase; Continuous gas fermentation; Genetic Enhancement - methods; Metabolic Engineering - methods; Metabolic Networks and Pathways - physiology; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Replicon for Gram-positives; Syngas
• ISSN: 1096-7176; 1096-7184
• DOI: 10.1016/j.ymben.2016.03.001

Expected depletion of oil and fossil resources urges the development of new alternative routes for the production of bulk chemicals and fuels beyond petroleum resources. In this study, the clostridial acetone pathway was used for the formation of acetone in the acetogenic bacterium Acetobacterium woodii. The acetone production operon (APO) containing the genes thlA (encoding thiolase A), ctfA/ctfB (encoding CoA transferase), and adc (encoding acetoacetate decarboxylase) from Clostridium acetobutylicum were cloned under the control of the thlA promoter into four vectors having different replicons for Gram-positives (pIP404, pBP1, pCB102, and pCD6). Stable replication was observed for all constructs. A. woodii [pJIR_actthlA] achieved the maximal acetone concentration under autotrophic conditions (15.2±3.4mM). Promoter sequences of the genes ackA from A. woodii and pta-ack from C. ljungdahlii were determined by primer extension (PEX) and cloned upstream of the APO. The highest acetone production in recombinant A. woodii cells was achieved using the promoters PthlA and Ppta-ack. Batch fermentations using A. woodii [pMTL84151_actthlA] in a bioreactor revealed that acetate concentration had an effect on the acetone production, due to the high Km value of the CoA transferase. In order to establish consistent acetate concentration within the bioreactor and to increase biomass, a continuous fermentation process for A. woodii was developed. Thus, acetone productivity of the strain A. woodii [pMTL84151_actthlA] was increased from 1.2mgL−1h−1 in bottle fermentation to 26.4mgL−1h−1 in continuous gas fermentation. •Acetone production from CO2 is possible by recombinant Acetobacterium woodii.•6 different acetone producing recombinant A. woodii strains were constructed.•DNA sequence of 2 promoters from acetogens were determined using PEX analysis.•3 plasmid replicons were identified that are suitable for replication in A. woodii.•Continuous gas fermentation increased acetone productivity up to 26.4mgL−1h−1.