Biotechnology for the Eucalyptus Biorefinery

Our work on the potential of oxidative biocatalysts in eucalypt biorefinery, including cellulose and biofuel production, is revised in the context of related studies. Organisms selectively degrading lignin from eucalypt wood are ideal models for these biotechnological applications. Among them, the white-rot fungus Ceriporiopsis subvermispora has been largely investigated for eucalypt "biopulping" (in combination with mechanical and chemical processes). Its recently sequenced genome provided us interesting information of the oxidative enzymes involved in wood decay. A large number of studies have been reported on these enzymes for "biobleaching" of eucalypt pulps, most of them focusing on the so-called laccase-mediator system. In collaboration with Novozymes, its industrial feasibility was demonstrated by using optimized doses of a commercial laccase and a phenolic mediator, with the double advantage of simultaneously removing lignin (responsible for pulp color) and lipids (causing pitch deposits). Recently, we showed that laccase, in the presence of mediators, is also able to remove lignin from eucalypt wood, in the absence of chemical pretreatments, resulting in increased saccharification and ethanol yields. In this case, lignin modification and selective removal were followed by a new procedure enabling 2D NMR analysis of the whole material at the gel stage. The availability of enzyme engineering tools, including directed evolution in addition to rational design, in combination with high-throughput methods and computational design is enabling us to optimize oxidative enzymes for efficient oxidation of different target substrates.