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- Continuous cultures of Clostridium acetobutylicum: culture stability and low-grade glycerol utilisationPublication . Andrade, José Carlos; Vasconcelos, IsabelContinuous cultures of two strains of Clostridium acetobutylicum were stable for over 70 d when grown on glucose/ glycerol mixtures. Butanol was the major fermentation end-product, accounting for 43 to 62% (w/w) of total products. Low-grade glycerol [65% (w/v) purity] could replace commercial glycerol [87% (w/v) purity], leading to a similar fermentation pattern: a butanol yield of 0.34 (mol/mol), a butanol productivity of 0.42 g l−1 h−1 and a 84% (w/w) glycerol consumptionwere attained when cultures were grown at pH 6 and D = 0.05 h−1; butanol accounted for 94% (w/w) of total solvents. These values are among the highest reported in literature for C. acetobutylicum simple chemostats.
- Metabolic engineering of clostridium acetobutylicum for the industrial production of 1,3-propanediol from glycerolPublication . González-Pajuelo, María; Meynial-Salles, Isabelle; Mendes, Filipa; Andrade, José Carlos; Vasconcelos, Isabel; Soucaille, PhilippeClostridium butyricum is to our knowledge the best natural 1,3-propanediol producer from glycerol and the only microorganism identified so far to use a coenzyme B12-independent glycerol dehydratase. However, to develop an economical process of 1,3-propanediol production, it would be necessary to improve the strain by a metabolic engineering approach. Unfortunately, no genetic tools are currently available for C. butyricum and all our efforts to develop them have been so far unsuccessful. To obtain a better ‘‘vitamin B12-free’’ biological process, we developed a metabolic engineering strategy with Clostridium acetobutylicum. The 1,3- propanediol pathway from C. butyricum was introduced on a plasmid in several mutants of C. acetobutylicum altered in product formation. The DG1(pSPD5) recombinant strain was the most efficient strain and was further characterized from a physiological and biotechnological point of view. Chemostat cultures of this strain grown on glucose alone produced only acids (acetate, butyrate and lactate) and a high level of hydrogen. In contrast, when glycerol was metabolized in chemostat culture, 1,3-propanediol became the major product, the specific rate of acid formation decreased and a very low level of hydrogen was observed. In a fed-batch culture, the DG1(pSPD5) strain was able to produce 1,3-propanediol at a higher concentration (1104mM) and productivity than the natural producer C. butyricum VPI 3266. Furthermore, this strain was also successfully used for very long term continuous production of 1,3-propanediol at high volumetric productivity (3 g L⁻¹ h⁻¹) and titer (788 mM).