Biosynthesis pathways of expanding carbon chains for producing advanced biofuels

Abstract

Because the thermodynamic property is closer to gasoline, advanced biofuels (C ≥ 6) are appealing for replacing non-renewable fossil fuels using biosynthesis method that has presented a promising approach. Synthesizing advanced biofuels (C ≥ 6), in general, requires the expansion of carbon chains from three carbon atoms to more than six carbon atoms. Despite some specific biosynthesis pathways that have been developed in recent years, adequate summary is still lacking on how to obtain an effective metabolic pathway. Review of biosynthesis pathways for expanding carbon chains will be conducive to selecting, optimizing and discovering novel synthetic route to obtain new advanced biofuels. Herein, we first highlighted challenges on expanding carbon chains, followed by presentation of two biosynthesis strategies and review of three different types of biosynthesis pathways of carbon chain expansion for synthesizing advanced biofuels. Finally, we provided an outlook for the introduction of gene-editing technology in the development of new biosynthesis pathways of carbon chain expansion.

Keywords: Advanced biofuels; CRISPR/Cas9; Carbon chains; Expanding.

Fig. 1

Development progress of bio-based small molecule compounds for replacing petroleum industry. With the development of bio-based small molecule compounds for substituting non-renewable gasoline, increasing length of carbon chain has been a hallmark of biosynthesis strategy. One of the major future developments of bio-based small molecule compounds for replacing fossil fuel is to exploit advanced biofuels with long carbon chains (C > 6) using appropriate engineering strategy

Fig. 2

Engineering strategy for producing short linear alcohols (C2–C5) based on the natural characteristics of microorganisms. Generally, synthesizing bio-based short linear alcohols (C2–C5) is based on the Ehrlich pathway and α-keto acid pathway of the microorganism itself. This engineering strategy is only possible to reach a maximum of five carbons, which cannot meet the necessary for extending carbon chain in generating non-natural bio-based small molecule compounds (C > 6) for substituting non-renewable gasoline. GAPDH: glyceraldehyde 3-phosphate dehydrogenase. PEP phosphoenolpyruvate. KDC 2-keto acid decarboxylases. ADH alcohol dehydrogenase. BAT2 aminotransferase. Double arrow show the oxidation–reduction pathway to biofuel

Fig. 3

The two biosynthesis strategies of expanding carbon chain in exploring new biosynthesis pathway. RM retrospective method. ED evolutionary design method. n reaction steps

Fig. 4

Biosynthesis pathway of expanding carbon chain for producing advanced biofuels using 2-keto acid intermediates. This biosynthesis pathway can be subdivided into three distinct sub-pathways (I, II, III), each of which results in a corresponding and structurally distinct advanced biofuel. a The I sub-pathway is that the carbon chain is extended via adding two carbon atoms (+ 2) with pyruvate as carbon extension unit; the key enzymes are acetohydroxy acid synthase (AHAS) and IlvIHCD. b The II sub-pathway is that the carbon chain is lengthened via adding one carbon atom (+ 1) with CoA-dependent molecule as carbon extension unit. Advanced biofuels with a branched chain are obtained via integrating the following natural biosynthetic pathways: α-keto acid-based pathway and CoA-dependent extension pathway. c Carbon chain extension of the Type III sub-pathway also occurs involving the addition of one carbon atom (+ 1) using CoA-dependent molecule as the carbon extension unit. Advanced biofuels without branched chains were obtained via utilizing this subtype pathway and conducting a condensation reaction with the overexpression of leuABCD. Green arrows represent using pyruvate as carbon extension unit. Blue arrows represent using CoA-dependent molecule as carbon extension unit. Red arrows represent exogenous decarboxylation and reduction. ilvA threonine deaminase; ilvC acetohydroxy acid isomeroreductase; ilvD dihydroxy acid dehydratase; ilvGM acetohydroxybutanoate synthase; kivd ketoisovalerate decarboxylase; leuA, 2-isopropylmalate synthase; leuB 3-isopropylmalate dehydrogenase; leuCD 2-isopropylmalate isomerase; thrA aspartate kinase, homoserine dehydrogenase; thrB homoserine kinase; thrC threonine synthase; Ac-CoA acetylCoA; ACP acyl carrier protein; CoA coenzyme A; OAA oxaloacetate; ADH2 alcohol dehydrogenase; alsS acetolactate synthase; asd aspartate semialdehyde dehydrogenase; aspC aspartate aminotransferase; cimA citramalate synthase. KIC:2-ketoisocaproate, precursor to l-leucine; KMV 2-keto-3-methylvalerate, precursor of l-isoleucine; PaaH1 β-ketothiolase (BKtB)-3-hydroxy-acyl-CoA dehydrogenase; Crt crotonase; Ter trans-enoyl-CoA reductase; adhE alcohol dehydrogenase. The red arrow shows the oxidation–reduction pathway to higher biofuel. The green arrow is the carbon chain extension pathway

Fig. 5

Biosynthesis pathway of expanding carbon chain for producing advanced biofuels using reverse β-oxidation intermediates. This biosynthesis pathway can use acetyl-CoA as a carbon chain elongation factor. Normal carbon chain extension in microorganisms occurs when (Cn + 2)-acyl-CoA molecules are broken down into acetyl-CoA and (Cn)-acetyl CoA molecules under the control of four genes (atoB, fadA, fadB, and fadE). However, chain elongation occurring through the addition of acetyl-CoA to another thioester due to β-oxidation cycle metabolic pathway is essentially reversible. Thus, advanced biofuels could be produced after one turn of this cycle. YqeF acetyl-CoA acetyltransferase; fadA ketoacyl-CoA thiolase; fadB hydroxyacyl-CoA dehydrogenase and enoyl-CoA hydratase; YdiO enoyl-CoA reductase; TES thioesterase; AcoAR acylCoA reductase; ADH alcohol dehydrogenase; aceEF-lpdA pyruvate dehydrogenase multi-enzyme complex; acetyl-CoA acetoacetyl-CoA transferase; atoB acetyl-CoA acyltransferase; atoE putative short-chain fatty acid transporter; dhaKLM PEP-dependent dihydroxyacetone kinase; fadA 3-ketoacyl-CoA thiolase; fadB enoyl-CoA hydratase/3-hydroxyaceyl-CoA dehydrogenase; fadD fatty acyl-CoA synthetase; fadE acyl-CoA dehydrogenase; fadL long-chain fatty acid outer membrane transporter; fbaA and fbaB: fructose-1,6-bisphosphate aldolase; fdhF formate dehydrogenase; galP galactose permease; gldA glycerol dehydrogenase; glk glucokinase; glpF glycerol MIP channel; hycB-1 hydrogenase; pflB pyruvate formate lyase; ptsHI-crr: ptsG glucose specific phosphoenolpyruvate phosphotransferase system; pykA and pykF pyruvate kinase; tpiA triose phosphate isomerase; xylA xyloseisomerase; xylB xylulokinase; xylE xylose MFS transporter; xylFGH: xylose ABC transporter. 2[H]: NAD(P)H = FADH2; DHA dihydroxyacetone, DHAP dihydroxyacetone-phosphate; Fructose-1: 6-BP fructose-1,6-bisphosphate. The red arrow shows the oxidation–reduction pathway to higher biofuel. The blue arrow is the carbon chain extension pathway

Fig. 6

Biosynthesis pathway to produce advanced biofuels of fatty alcohols using fatty acid intermediates. Advanced biofuels: fatty alcohols product from carbon chain extension of fatty acid synthesis. Mechanistically, carbon chain extension relies on acetyl-CoA as the primary carbon source in the fatty acid synthetic pathway via ATP-dependent carboxylation into malonyl-CoA. Acyl units are then provided for carbon elongation by cycles of decarboxylative addition of malonyl-CoA. accABCD acetyl-CoA carboxylase; acrI acyl-CoA reductase; asd aspartate semialdehyde dehydrogenase; atfA acyltransferase; fabA hydroxydecanoyl-ACP dehydrase; fabB ketoacyl-ACP synthase I; fabD malonyl-CoA-ACP transacylase; fabF ketoacyl-ACP synthase II; fabG ketoacyl-ACP reductase; fabH ketoacyl-ACP synthase III; fabI enoyl-ACP reductase; fabZ hydroxyacyl-ACP dehydratase; CER4 fatty acyl-CoA reductase acyl-CoA synthetase; FAEE fatty acid ethyl ester; Ac-CoA acetylCoA; ACP acyl carrier protein; CoA coenzyme A

Fig. 7

Biosynthesis pathway to produce advanced biofuels of alkanes and alkenes using fatty acid intermediates. Increasing microbial synthesis of alka(e)nes via carbon chain extension of five different fatty acid synthesis pathways that can convert free fatty acids or fatty acid derivatives into alka(e)nes, classified as “elongation–decarboxylation” and “head-to-head condensation” biosynthetic metabolic approaches, including AAR-ADO, HEAD-TO-HEAD, OLeTJE, OLS, and CAR/FAR-ADO pathways. The elongation–decarboxylation approach is based on the use of acyl-coenzyme A (CoA) from a fatty acid synthesis pathway as an extension factor for carbon chain elongation via the circular addition of a two-carbon unit, conversion of malonyl-CoA, and subsequent decarboxylation to produce chain alka(e)nes. The second approach is head-to-head condensation, involving the conjugation of two fatty acid derivatives to a carboxylic acid via a Claisen condensation. The odd chain alka(e)ne is formed by decarboxylation and decarbonylation. ACP acyl–acyl carrier protein; ACC acetyl-CoA carboxylase; FabD malonyl-CoA ACP transacylase; FabH β-keto-acyl-ACP synthase III; FabB β-keto-acyl-ACP synthase I; FabG β-keto-acyl-ACP reductase; FabZ: β-hydroxyacyl-ACP dehydratase; FabI enoyl-acyl-ACP reductase; TE thioesterase; FadD acyl-CoA synthase; AAR acyl-ACP reductase; ADO: aldehyde-deformylating oxygenase; OleABCD: a four protein families for long-chain olefin biosynthesis; (OleTJE): fatty acid decarboxylase, a cytochrome P450 enzyme that reduces fatty acids to alkenes; CAR: carboxylic acid reductase; Sfp: A phosphopantetheinyl transferase; FAR: fatty acid reductase; Ols: a type I polyketide synthases for α-olefin biosynthesis. MVA indicates mevalonic pathway for isoprenoid biosynthesis. MEP indicates the methylerythritol phosphate pathway. Red, purple, blue fonts show the oxidation–reduction pathway to higher biofuel

Fig. 8

The complex relationship in different biosynthesis pathways of expanding carbon chain. Every one-carbon chain extension generates a global net effect per catalytic cycle regardless of whether the pathway involving 2-keto acid intermediate (“ + 1” or “ + 2” pathway) or a biosynthesis pathway using fatty acid intermediates and reverses β-oxidation. A mixture containing various advanced biofuels namely, longer, branched or unbranched carbon chains (e.g., alcohols) is capable of producing via the per addition of one carbon. Gly-3-P: glyceraldehyde-3-phosphate; PEP phosphoenolpyruvate; Pv pyruvate; MC malonyl-CoA; MA malonyl-ACP; AC acyl-CoA