Identifying, characterising and engineering fungal plant cell wall degrading enzymes for enhanced biocatalysts in biofuel production

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BBSRC Theme: Industrial Biotechnology and BioenergyFungal and oomycete genomes are a rich source of carbohydrate processing enzymes that are capable of plant cell wall degradation. Exploitation of these enzymes is attractive given they have the capability of converting waste lignocellulosic biomass from plants into simpler sugars, which can be utilised in biofuel production.We will focus on fungal/oomycete species including Rhynchosporium spp., Magnaporthe spp., and Phytophthora spp. These species possess a range of carbohydrate processing specificities with potential in biocatalysis. The first part of the project will involve biomining the genomes of a range of fungal/oomycete species to identify carbohydrate degrading enzymes. The focus will be on enzyme families that have potential for natural and engineered substrate diversity, and which present a promising basis for further engineering.Following this, the genes encoding a number of these enzymes will be cloned and over-expressed in Escherichia coli, and their activity and specificity against a range of carbohydrate substrates investigated. In addition, the structures of the enzymes will be determined using X-ray crystallography. These structural and functional insights will be used to identify candidates to take forward for engineering, where we will use a rational approach to design mutants, including those where the natural sequence diversity of related enzymes can be incorporated, to explore the potential for enhanced activity and/or a wider substrate specificity. Engineering may require combinations of mutants and iterative cycles in order to obtain enzymes that can be applied in biocatalytic applications.