Genetic engineering of algae for enhanced biofuel production

Abstract

There are currently intensive global research efforts aimed at increasing and modifying the accumulation of lipids, alcohols, hydrocarbons, polysaccharides, and other energy storage compounds in photosynthetic organisms, yeast, and bacteria through genetic engineering. Many improvements have been realized, including increased lipid and carbohydrate production, improved H(2) yields, and the diversion of central metabolic intermediates into fungible biofuels. Photosynthetic microorganisms are attracting considerable interest within these efforts due to their relatively high photosynthetic conversion efficiencies, diverse metabolic capabilities, superior growth rates, and ability to store or secrete energy-rich hydrocarbons. Relative to cyanobacteria, eukaryotic microalgae possess several unique metabolic attributes of relevance to biofuel production, including the accumulation of significant quantities of triacylglycerol; the synthesis of storage starch (amylopectin and amylose), which is similar to that found in higher plants; and the ability to efficiently couple photosynthetic electron transport to H(2) production. Although the application of genetic engineering to improve energy production phenotypes in eukaryotic microalgae is in its infancy, significant advances in the development of genetic manipulation tools have recently been achieved with microalgal model systems and are being used to manipulate central carbon metabolism in these organisms. It is likely that many of these advances can be extended to industrially relevant organisms. This review is focused on potential avenues of genetic engineering that may be undertaken in order to improve microalgae as a biofuel platform for the production of biohydrogen, starch-derived alcohols, diesel fuel surrogates, and/or alkanes.

Fig. 1.

Microalgal metabolic pathways that can be leveraged for biofuel production. ER, endoplasmic reticulum.

Fig. 2.

Simplified overview of the metabolites and representative pathways in microalgal lipid biosynthesis shown in black and enzymes shown in red. Free fatty acids are synthesized in the chloroplast, while TAGs may be assembled at the ER. ACCase, acetyl-CoA carboxylase; ACP, acyl carrier protein; CoA, coenzyme A; DAGAT, diacylglycerol acyltransferase; DHAP, dihydroxyacetone phosphate; ENR, enoyl-ACP reductase; FAT, fatty acyl-ACP thioesterase; G3PDH, gycerol-3-phosphate dehydrogenase; GPAT, glycerol-3-phosphate acyltransferase; HD, 3-hydroxyacyl-ACP dehydratase; KAR, 3-ketoacyl-ACP reductase; KAS, 3-ketoacyl-ACP synthase; LPAAT, lyso-phosphatidic acid acyltransferase; LPAT, lyso-phosphatidylcholine acyltransferase; MAT, malonyl-CoA:ACP transacylase; PDH, pyruvate dehydrogenase complex; TAG, triacylglycerols.

Fig. 3.

Starch metabolism in green microalgae. The metabolites and simplified representative pathways in microalgal starch metabolism are shown in black, and enzymes are shown in red. Glucans are added to the water soluble polysaccharide (WSP) by α-1,4 glycosidic linkages (WSP₁) until a branching enzyme highly branches the ends (WSP₂). Some of these branches are trimmed (WSP₃), and this process is repeated until a starch granule is formed. Phosphorolytic [Starch-(P)_n] and hydrolytic degradation pathways are shown. αAMY, α-amylase; AGPase, ADP-glucose pyrophosphorylase; βAMY, β-amylases; BE, branching enzymes; DBE, debranching enzymes; DPE, disproportionating enzyme (1 and 2) α-1,4 glucanotransferase; Glc, glucose; GWD, glucan-water dikinases; ISA, isoamylases; MEX1, maltose transporter; MOS, malto-oligosaccharides; PGM, plastidial phosphoglucomutase; P, phosphate; P_i, inorganic phosphate; PP_i, pyrophosphate; SP, starch phosphorylases; SS, starch synthases.