Synthetic Biology Applied to Carbon Conservative and Carbon Dioxide Recycling Pathways

Abstract

The global warming conjugated with our reliance to petrol derived processes and products have raised strong concern about the future of our planet, asking urgently to find sustainable substitute solutions to decrease this reliance and annihilate this climate change mainly due to excess of CO₂ emission. In this regard, the exploitation of microorganisms as microbial cell factories able to convert non-edible but renewable carbon sources into biofuels and commodity chemicals appears as an attractive solution. However, there is still a long way to go to make this solution economically viable and to introduce the use of microorganisms as one of the motor of the forthcoming bio-based economy. In this review, we address a scientific issue that must be challenged in order to improve the value of microbial organisms as cell factories. This issue is related to the capability of microbial systems to optimize carbon conservation during their metabolic processes. This initiative, which can be addressed nowadays using the advances in Synthetic Biology, should lead to an increase in products yield per carbon assimilated which is a key performance indice in biotechnological processes, as well as to indirectly contribute to a reduction of CO₂ emission.

Keywords: bio-based products; carbon dioxide; chemicals; metabolic engineering; microbial physiology; synthetic biology.

Figure 1

The scheme of non–oxidative glycolysis (NOG), Embden-Pentose-Bifido (EP-Bifido), and Glycoptimus synthetic pathways. Enzymes abbreviation are Fxpk, F6P/Xu5P-phosphoketolase; Tal, transaldolase; Tkt, transketolase; Fbp, fructose-1,6- bisphosphatase; Fba, fructose1,6 bisphosphate aldolase; tpi, triose phosphate isomerase; Rpe, ribulose 5-P epimerase; Rpi, ribose 5-P isomerase; Kdsd, arabinose-5-P isomerase; Fsa, fructose-6P aldolase; Alda, glycolaldehyde dehydrogenase. Metabolites: F6P, fructose-6P; PEP, phosphoenolpyruvate; Pyr, pyruvate; E4P, erythrose-4P; S7P, sedoheptulose-7P; R5P, ribose-5P; Ru5P, ribulose-5P; X5P, xylulose-5P; GAP, glyceraldehyde-3P; DHAP, dihydroxyacetone-P; FBP, fructose-1,6-bisphosphate; AcP, acetyl-Pi, AcCoA, acetyl-CoA.

Figure 2

The methanol condensation cycle (MCC) for assimilation of methanol into higher alcohol with maximal carbon conservation. The MCC is a combination of the RuMP with part of the NOG allowing to avoid pyruvate decarboxylation and to bypass ATP dependency. It results in the net production of acetyl-CoA (AcCoA) from 2 methanol. Here, it is represented the metabolic pathway leading to the production of n-butanol which requires 4 methanol. Enzymes abbreviations are: Medh, NAD⁺-dependent methanol dehydrogenase; Hps, hexulose-6P synthase; Phi, phosphohexuloseisomerase; Tkt, transketolase, Fxkp, F6P/Xu5P phosphoketolase; Tal, transaldolase; Rpe, ribulose-5-P epimerase; Rpi, ribose-5-isomerase. Abbreviation for metabolites as in Figure 1.

Figure 3

Scheme of the reversal of glyxoylate shunt (rGS) and maly-CoA Glycerate (MGC) cycling pathway for the production of acetyl-CoA from sugars with maximal carbon conservation. Enzymes abbreviation are: Ppc, PEP carboxylase; Mdh, malate dehydrogenase; Fum, fumarase; Frd, fumarate reductase; AceA, isocitrate lyase; Aco, aconitase; Acl, acyl-CoA lyase: Mtk, malate thiokinase; Mcl, malyl-CoA lyase; Gdh, glycolate dehydrogenase; Gcl, glyoxylate carboxyligase; Tsr, tartronate semialdehyde reductase; Glk, glycerate kinase; Eno, enolase.

Figure 4

Scheme of Malonyl-coA-Oxaloacetate- Glyoxylate (MOG), Modified Serine (MSer) and 4-hydroxy-2-oxo-butyrate (HOB) cycling pathway employing the efficient PEP carboxylase to capture Co₂ into either product or biomass. Enzymes abbreviation are: Ppc, PEP carboxylase; Mdh, malate dehydrogenase; Mtk, malate thiokinase; Mcl, malyl -CoA lyase; Mct, malonyl-CoA acetyl transferase; Mcr, malonyl CoA reductase; Bapta, β-alanine: pyruvate transaminase; Aam, alanine 2,3 aminomutase; Pps, PEP synthase; Ta, transaminase; Ask, aspartate kinase; Asd, aspartyl-Pi semialdehyde dehydrogenase; HmdH, homoserine dehydrogenase; Hmtf, hydroxymethyl transferase.

Figure 5

Engineering sugars fermentation with Calvin-cycle enzymes for fixing in-situ CO₂ to improve bioethanol yield in yeast. In green are shown the Calvin-cycle enzymes corresponding to RuBisCO and phosphoribulose-5-P kinase (PRK). The expression of this Calvin-cycle enzymes enables the use of CO₂ as alternative electron acceptor for reoxidation of NADH, thereafter reducing glycerol production under anaerobic condition. Enzymes: RuBisCO, ribulose 1,5 bisphosphate carboxylase; PRK, phosphoribulose 5-P kinase; Tkt, transketolase; Tal, transaldolase. Metabolites abbreviation, as in Figure 1 except: 1,3-DPG, 1,3 diphosphoglycerate; 3-PG, 3-phosphoglycerate; 2-PG, 2-phosphoglycerate; G3P, glycerol-3-P.