• Open Access

Biosynthesis and incorporation of side-chain-truncated lignin monomers to reduce lignin polymerization and enhance saccharification

Authors

  • Aymerick Eudes,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
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  • Anthe George,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Sandia National Laboratory, Livermore, CA, USA
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  • Purba Mukerjee,

    1. Department of Biochemistry, The Wisconsin Bioenergy Initiative, and the DOE Great Lakes Bioenergy Research Center, University of Madison, Madison, WI, USA
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  • Jin S. Kim,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
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  • Brigitte Pollet,

    1. Institut Jean Pierre Bourgin, INRA-AgroParis Tech, UMR1318, Versailles, France
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  • Peter I. Benke,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
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  • Fan Yang,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
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  • Prajakta Mitra,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
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  • Lan Sun,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Sandia National Laboratory, Livermore, CA, USA
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  • Özgül P. Çetinkol,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Sandia National Laboratory, Livermore, CA, USA
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    • Present address: Chemistry Division, Doğuş University, Istanbul, Turkey.

  • Salem Chabout,

    1. Institut Jean Pierre Bourgin, INRA-AgroParis Tech, UMR1318, Versailles, France
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  • Grégory Mouille,

    1. Institut Jean Pierre Bourgin, INRA-AgroParis Tech, UMR1318, Versailles, France
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  • Ludivine Soubigou-Taconnat,

    1. UMR INRA 1165 CNRS 8114, Recherche en Génomique Végétale, UEVE, Evry Cedex, France
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  • Sandrine Balzergue,

    1. UMR INRA 1165 CNRS 8114, Recherche en Génomique Végétale, UEVE, Evry Cedex, France
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  • Seema Singh,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Sandia National Laboratory, Livermore, CA, USA
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  • Bradley M. Holmes,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Sandia National Laboratory, Livermore, CA, USA
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  • Aindrila Mukhopadhyay,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
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  • Jay D. Keasling,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
    3. Departments of Bioengineering, Chemical & Biomolecular Engineering, University of California, Berkeley, CA, USA
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  • Blake A. Simmons,

    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Sandia National Laboratory, Livermore, CA, USA
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  • Catherine Lapierre,

    1. Institut Jean Pierre Bourgin, INRA-AgroParis Tech, UMR1318, Versailles, France
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  • John Ralph,

    1. Department of Biochemistry, The Wisconsin Bioenergy Initiative, and the DOE Great Lakes Bioenergy Research Center, University of Madison, Madison, WI, USA
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  • Dominique Loqué

    Corresponding author
    1. Joint BioEnergy Institute, EmeryStation East, Emeryville, CA, USA
    2. Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
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(Tel +1 510 486 7332; fax +1 510 486 4252; email dloque@lbl.gov)

Summary

Lignocellulosic biomass is utilized as a renewable feedstock in various agro-industrial activities. Lignin is an aromatic, hydrophobic and mildly branched polymer integrally associated with polysaccharides within the biomass, which negatively affects their extraction and hydrolysis during industrial processing. Engineering the monomer composition of lignins offers an attractive option towards new lignins with reduced recalcitrance. The presented work describes a new strategy developed in Arabidopsis for the overproduction of rare lignin monomers to reduce lignin polymerization degree (DP). Biosynthesis of these ‘DP reducers’ is achieved by expressing a bacterial hydroxycinnamoyl-CoA hydratase-lyase (HCHL) in lignifying tissues of Arabidopsis inflorescence stems. HCHL cleaves the propanoid side-chain of hydroxycinnamoyl-CoA lignin precursors to produce the corresponding hydroxybenzaldehydes so that plant stems expressing HCHL accumulate in their cell wall higher amounts of hydroxybenzaldehyde and hydroxybenzoate derivatives. Engineered plants with intermediate HCHL activity levels show no reduction in total lignin, sugar content or biomass yield compared with wild-type plants. However, cell wall characterization of extract-free stems by thioacidolysis and by 2D-NMR revealed an increased amount of unusual C6C1 lignin monomers most likely linked with lignin as end-groups. Moreover the analysis of lignin isolated from these plants using size-exclusion chromatography revealed a reduced molecular weight. Furthermore, these engineered lines show saccharification improvement of pretreated stem cell walls. Therefore, we conclude that enhancing the biosynthesis and incorporation of C6C1 monomers (‘DP reducers’) into lignin polymers represents a promising strategy to reduce lignin DP and to decrease cell wall recalcitrance to enzymatic hydrolysis.

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