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Genetic Modification to Improve Bioconversion of Aspen Wood to Ethanol

“Woody biomass derived from the transgenic aspens shows a 62% increase in the release of simple sugars and up to a 49% increase in the yield of ethanol”

Wood is one of the oldest fuel sources known to man but its use declined with the introduction of more efficient fossil fuels. However, with the depletion of fossil fuel stores and the need to reduce emissions it has been making a comeback. As the need for clean, renewable fuel sources increases, ways to improve the efficiency of biofuel production are continually being sought.

The ongoing supply of wood from managed forests make it a particularly attractive biofuel. In addition to being burnt to produce steam to create electricity, wood can also be used to produce the alcohol bioethanol, which can be used to fuel vehicles. Bioethanol is made by fermenting plant materials, including wood.

Unfortunately, the lignin in plant cell walls that gives wood its strength makes processing woody biomass into biofuels and bio-based chemicals more difficult. The cellulose of wood is impregnated with lignin, which forms a hydrophobic barrier protecting the wood. Since 20% of aspen wood is lignin, this significantly limits its conversion to fermentable sugars (saccharification) during the biofuel production process since it is not easily reached by digestive enzymes.

It has been reported that reducing the lignin content of plant cell walls can improve the efficiency of biomass conversion, whereby lowering costs. However, there are concerns that such modifications could reduce biomass yield.

A recent study investigated the effects of preventing lignin monomeric precursors from being incorporated into the lignin polymer through genetic engineering of aspen trees. The modified trees were engineered to overproduce the enzyme monolignol 4-O-methyltransferase (MOMT4). This enzyme catalyzes the methylation of lignin precursors, thereby disrupting lignin formation. Nuclear magnetic resonance imaging (acquired on a Bruker Biospin Avance 700MHz spectrometer) confirmed that the lignin content and structure in the resulting trees was significantly different from the native aspens (Figure 1). Although the overall lignin content was not markedly lower in the modified aspen trees, there was a significant reduction in the number of S-type lignin subunits in the cell walls. Thus, modification of the MOMT4 enzyme had impaired the trees’ ability to incorporate the S-lignin monomer during cell wall manufacture. Instead, they appear to have compensated for this deficiency by incorporating more cellulose. The total cellulose content of cell walls was 12% higher in the modified trees than in the native trees. The observed cell wall structural changes were not associated with a reduction in growth or biomass production of the modified trees.

Furthermore, compared with the native trees, woody biomass obtained from the aspen trees with impaired lignin synthesis yielded 62% more simple sugars and up to 49% more ethanol when subjected to enzymatic digestion and yeast-mediated fermentation. In addition, the rate at which saccharification and bioconversion occurred was higher for the modified wood compared with the native wood.

Such impairment of lignin synthesis could thus provide a useful means for improving the efficiency of woody biomass processing for bio-based applications.

Figure 1. The effect of expression of MOMT4 on lignin content and composition in transgenic aspens

aspen_wood

(a) Three-month-old hybrid aspens of control (left) and three MOMT4 independent transgenic lines (right). (b,c) Phloroglucinol-HCl staining of the stem cross-sections of control (b) and MOMT4-0 transgenic line (c). (d,e) Ma¨ule staining of the stem cross-sections of control (d) and MOMT4-0 transgenics (e). Scale bars, 1 mm. (f) Acetyl bromide total lignin content in the cell walls of control and MOMT4 transgenic aspen stems. (g) The monomers released by thioacidolysis from the stem cell walls of MOMT4 transgenic aspens; S, syringyl; G, guaiacyl; H, p-hydroxyphenyl; CWR, cell wall residues; Ctrl., control. Data in f,g represent mean±s.e.with three biological replicates (each with three technical repeats) for the control and three technical repeats for the individual transgenic lines. ** Indicates significant difference of lignin content (f) or S-monomer (g) compared to the control with Po0.01 (Student’s t-test).

Cai Y, et al. Enhancing digestibility and ethanol yield of Populus wood via expression of an engineered monolignol 4-O-methyltransferase. Nat Commun. 2016 Jun 28;7:11989. doi: 10.1038/ncomms11989 (2016).

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