Harnessing invertebrate wood digestion systems

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Lignocellulosic biomass represents the greatest reserve of fixed carbon in the biosphere and its degradation is an important component of the global carbon cycle. Lignocellulose, in the form of crop residues and the biological fraction of municipal solid waste, also represents a potential low-carbon replacement for petroleum for the production of sustainable fuels, chemicals and materials. Lignocellulose is a rich potential source of sugars and phenolics as it is primarily comprised of polysaccharides and lignin. These can serve as a source of dietary nutrition for organisms that feed on it, or as platform chemicals for industry, but in both cases, obtaining these compounds is challenging due to the recalcitrance of lignocellulose. We are studying the digestive systems of invertebrate animals specialised to live on a diet of lignocellulosic biomass. These studies can help us to understand important environmental components of the global carbon cycle, as well as providing us with new understanding and tools to improve the industrial exploitation of lignocellulosic biomass.The organisms we study include two species of crustacean woodborers (Limnoria tripunctata, and Chelura terebrans), a bivalve molluscan shipworm (Lyrodus pedicellatus) and an insect detritivore (Thermobia domestica). We used transcriptomic and proteomic analyses to study the genes and proteins expressed in the digestive system of these animals in order to identify the major digestive enzymes that they use. Our studies reveal clear similarities and novel features of the digestive proteome of the different species. Our work has already revealed a number of novel enzyme classes from invertebrate lignocellulose degraders. These include: the first report of processive cellobiohydrolases from animals, a new class of lytic polysaccharide monooxygenases (and the first reported in animals), and a new class of lignin active enzymes that speed up the digestion of wood by cellulases. Our studies show that the animals focus on digesting the cellulosic component of lignocellulose as this is made of polymers of glucose, which is easy to metabolise. However, in order to access the cellulose they have to disrupt the lignin and hemicellulose components that encase the cellulose. We have recently used 2-dimensional NMR studies of faecal residues from Limnoria and Lyrodus to identify major changes in wood composition during digestion. These reveal that there is extensive removal of the glucuronic acid and acetyl side chains of hemicellulose during digestion. Recent work from other groups suggests that the glucuronic acid side chains of xylans (a major hemicellulose) provide a link to the lignin, and that the acetyl esters help stabilise the associations between xylans and cellulose.Our proposed work is to identify and characterise some of the, as-yet uncharacterised, major activities from our invertebrate systems. These include two families of glycosyls hydrolases (GHs) found in all four digestive systems (GH9s and GH30s) and which we have now been able to produce in a recombinant form, the lignin-modifying enzymes of Lyrodus (now available in recombinant forms), and acetyl esterases and glucuronidases from Lyrodus and Limnoria. We will use high-throughput robotic facilities in our lab to test combinations of enzymes from our collection for their ability to synergise the deconstruction of lignocellulosic and cellulosic substrates. We will also assess the ability of our enzymes to enhance the activity of commercial cellulase preparations and work with partners from the enzyme industry to develop ways to exploit our discoveries.