Clostridium ljungdahlii represents a microbial production platform based on syngas

Abstract

Clostridium ljungdahlii is an anaerobic homoacetogen, able to ferment sugars, other organic compounds, or CO(2)/H(2) and synthesis gas (CO/H(2)). The latter feature makes it an interesting microbe for the biotech industry, as important bulk chemicals and proteins can be produced at the expense of CO(2), thus combining industrial needs with sustained reduction of CO and CO(2) in the atmosphere. Sequencing the complete genome of C. ljungdahlii revealed that it comprises 4,630,065 bp and is one of the largest clostridial genomes known to date. Experimental data and in silico comparisons revealed a third mode of anaerobic homoacetogenic metabolism. Unlike other organisms such as Moorella thermoacetica or Acetobacterium woodii, neither cytochromes nor sodium ions are involved in energy generation. Instead, an Rnf system is present, by which proton translocation can be performed. An electroporation procedure has been developed to transform the organism with plasmids bearing heterologous genes for butanol production. Successful expression of these genes could be demonstrated, leading to formation of the biofuel. Thus, C. ljungdahlii can be used as a unique microbial production platform based on synthesis gas and carbon dioxide/hydrogen mixtures.

Fig. 1.

Scheme of basic metabolic pathways and energy conservation in C. ljungdahlii when growing heterotrophically on sugars (hexoses or pentoses) or autotrophically on gases (CO or CO₂ + H₂). Glycolysis and pentose phosphate pathway for heterotrophic growth are underlaid in gray as well as the Wood–Ljungdahl pathway for autotrophic growth. The white boxes represent substrates and the dark gray boxes with white letters represent products. Single reactions do not represent stoichiometric fermentation balances. Ack, acetate kinase; ACS, acetyl-CoA synthase; AdhE, butyraldehyde/butanol dehydrogenase E; AOR, aldehyde oxidoreductase; Co-FeS-P, corrinoid iron-sulfur protein; Etf, electron-transferring flavoprotein; Fd, ferredoxin; GAP, glyceraldehyde-3-phosphate; PFOR, pyruvate:ferredoxin-oxidoreductase; Pta, phosphotransacetylase; PTS, PEP-dependent phosphotransferase system; THF, tetrahydrofolate.

Fig. 2.

Rnf complex in C. ljungdahlii.

Fig. 3.

Growth experiments with C. ljungdahlii wild type, C. ljungdahlii (pIMP1), and C. ljungdahlii (pSOBP_ptb) in 1,000-mL flasks with 200 mL PETC media and fructose or 0.8 bar synthesis gas. OD_600nm (circles) and concentration of acetate (triangles up), ethanol (squares), butanol (triangles down), and butyrate (diamonds) were measured in three different cultures (black, dark gray, and light gray).

Fig. 4.

Northern blots of recombinant C. ljungdahlii (pSOBP_ptb). Probes used: thlA and adhE from C. acetobutylicum and ack from C. ljungdahlii. Samples were taken from exponential (exp.) and stationary (stat.) growth phases.