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Genetically Altering Lignin Composition to Improve Biomass Processing

“the C2-Idf mutant produces highly reduced levels of apigenin- and tricin-related flavonoids, resulting in a strongly reduced incorporation of tricin into the lignin polymer.”

Second generation biofuels are advanced biofuels that have been developed to counteract the limitations of first-generation biofuels, which do not serve as sustainable substitutions for oil products or meet the targets set for climate change.

One of the most promising first-generation biofuels is lignocellulosic biomass, a bio-renewable, carbon-neutral plant biomass that can be produced from agricultural crop residues and inedible plant tissue.

Lignocellulosic biomass is mainly made up of plant secondary cell walls, which are composed of cellulose, hellicellluose and lignin. Cellulose has large reservoirs of energy, thereby offering great potential for conversion to biofuels.

Fermentable sugars are generated from cellulose by enzymatic degradation, but this process is hindered by the complex structure and make-up of the plant cell wall, a feature that is referred to as cell wall recalcitrance. In particular, the presence of the phenolic heteropolymer lignin limits this enzymatic digestibility by immobilizing hydrolytic enzymes and preventing enzymes from accessing polysaccharides.

In dicotyeldons, a class of flowering plants, the lignin polymer is initiated by monolignol dimerization, whereas in monocotyledons such as grass, tricin is incorporated into the native lignin polymer.

The flux towards flavonoid biosynthesis and therefore tricin is controlled by an enzyme called chalcone synthase (CHS) and in the monocotyledon maize, the main gene that codes for this enzyme is Colorless2 (C2).

In a recent study conducted by Wout Boerjan and colleagues, the researchers investigated a maize mutant that has reduced tricin levels due to the presence of an inhibitory mutation called C2-Idf (inhibitor diffuse), that silences expression of the C2 gene.

The team investigated the effects that an altered tricin level had on lignin structure and cell wall recalcitrance using phenolic profiling, saccharification assays and nuclear magnetic resonance imaging of the maize. For the NMR experiments, Bruker BioSpin’s AVANCE 700-MHz spectrometer was used to obtain NMR spectra.

Using NMR spectroscopy in this way allowed Boerjan and team to discover that the level of tricin-related flavonoids was highly reduced in the C2-Idf mutant, which resulted in strongly reduced incorporation of tricin into the lignin polymer. Analysis of unit enrichment in the lignin confirmed that tricin initiates lignin chains.

Furthermore, the absence of tricin led to more monolignol dimerization reactions and the effects this had on lignin structure impacted the saccharification efficiency of the mutant maize leaves.

The researchers say the findings are instructive for lignin engineering strategies that could improve biomass processing and biochemical production.

For example, incorporating tricin into dicotyledon lignins that do not otherwise produce tricin, or genetically engineering the overproduction of tricin in monocotyledons, could serve as potential strategies to test the direct impact tricin levels have on lignocellulose recalcitrance.

Contact Bruker for more information about the AVANCE 700-MHz spectrometer.

Reference:

  • Boerjan, W et al. Silencing CHALCONE SYNTHASE in Maize Impedes the Incorporation of Tricin into Lignin and Increases Lignin Content. Plant Physiology 2017;173:998–1016