Metabolic engineering of Clostridium acetobutylicum for the industrial production of 1,3-propanediol from glycerol

Abstract

Clostridium 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 (1104 mM) 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).

Anaerobic fermentation of glycerol: a platform for renewable fuels and chemicals

“…with glycerol as the sole carbon source has resulted in 1,3-PDO titers greater than 60 g/l from both isolated and engineered strains of K. pneumoniae [31][32][33], C. butyricum [31,[34][35][36][37], and recombinant Clostridium acetobutylicum [34,38]. Due in part to the large number of studies and interest in the conversion of glycerol into 1,3-PDO, this topic has also been extensively reviewed [20,39].…”

Continuous flow transformations of glycerol to valuable products: an overview

“…Clostridium acetobutylicum) [22]. In terms of chemical production of 1,3-PDO, an aqueous-phase glycerol degradation protocol was reported to employ a Pt/WO 3 residence time) [23].…”

Intensive Production of Carboxylic Acids Using <em>Cl. butyricum</em> in a Membrane Bioreactor (MBR)

“…As carbon input the carbon molecules contained in the carbohydrate and nitrogen feed source, in this case glucose and yeast extract, were considered while cellular biomass, carbon dioxide and acetic and butyric acid are held as carbon output. These calculation were made taking into consideration the following assumptions: a) cell biomass (CH1.77O0.49 N0.24 ) contains 50 wt% carbon [39] , b) yeast extract which is mainly comprised of the water-soluble components of the yeast cell, the composition of which is primarily amino acids, peptides, carbohydrates, and salt, is assumed containing at least 50 wt% carbon [40,41] that is consumed fully c) during Cl. butyricum metabolism , for each mole of acetic acid produced, 1 mole of CO2 is generated, while for each mole of butyric acid 2 moles of CO2 are produced (Fig.…”