Caractérisation biochimique, structurale et fonctionnelle de aryl etherases – Etherase

The first objective of this proposal is to characterize a new family of GST related to aryl-etherases (GTEs). From literature data and previous expression data performed in the UMR1136 unit, it appears that GTE family is particularly extended in ligninolytic fungi, suggesting a function of these enzymes in lignin mineralization. Studies concerning the distribution and the function of these enzymes will give new functional insights on the microbial community involved in biomass degradation. Besides the biochemical and structural characterization of different enzymes arising from model organisms (P. chrysosporium, P. placenta), presence and expression of GTEs will be tested both in fungal collections and in specific environmental samples chosen for their interest. We expect indeed to develop new biomarkers in particular for the wood industry. Wood decay is one of the most significant problems facing the wood products industry, limiting wood product utilization in adverse environments and costing millions of Euros annually for decay prevention and replacement of woody structural elements. Detecting and monitoring decay in cellulose and wood products ranging from bridges, to private homes and public buildings remains a problem, particularly where wood is in ground contact or exposed to moisture. Optimal methods for the detection of decay fungi in wood and cellulose insulation products have not, however, been developed. The development of a reliable assay for the early detection of decay in wood is also needed. Neutral markers-based molecular tools are being developed to detect fungi with special focus on some species but a powerful large spectrum identification tool is still needed. Moreover, species identification is not enough to evaluate the destruction potential of detected fungi. Since many species cannot be cultivated and cultivation take time, fast and easy-to-use molecular or/and enzymatic diagnostic tools for ligno/cellulolytic activity would be of high interest for decay fungi in buildings as well as for industrial fungi lignin removal (pulp and paper industry, ethanol production and other bio-refinery activity). The abundance and the specificity of GTEs in microorganisms involved in lignin degradation (suggest that aryl etherases should be good candidates as functional markers of wood degradation. The second objective of this project is to characterize extracellular enzymes able to hydrolyze -aryl ether linkages of lignin related compounds. This approach is quite original since only one fungal extracellular enzyme has been previously partially characterized from a Chaetomium-related fungus. In view of rising prices of crude oil due to increasing fuel demands, the need for alternative sources of bioenergy is expected to increase sharply in the coming years. The 'first-generation' biofuels made from starch and sugar appear unsustainable because of the potential stress that their production places on food commodities. Second-generation biofuels, produced from cheap and abundant plant biomass, are seen as the most attractive solution to this problem. At present, the production of such fuels is not cost-effective because there are a number of technical barriers that need to be overcome before their potential can be realized and particularly the need to overcome the saccharification barrier due mainly to the polysaccharides accessibility in plant cell wall. There is widespread agreement that there remains great potential for the discovery of novel enzymes to be used in tackling the saccharification barrier. Such enzymes may be employed directly during processing in vitro to overcome the recalcitrance of lignocellulose to cellulases. Clearly, enzymes able to cleave specifically the major linkages found in lignins are very good candidates, in contrast to non-specific oxidative systems. Indeed, lignin-degrading systems characterized so far involved free radical mechanisms based upon radicals generated mainly by complex enzymatic systems as lignin peroxidases, manganese peroxidases, different oxidases'. The complexity of these systems involving a high number of different haem-containing enzymes and requiring the continuous production of hydrogen peroxide limits considerably the use of such systems at the industrial level. As described above, strong evidences suggest that, in nature, '-aryl etherases are involved in the degradation of lignin-related compounds released by the action of wood-decay fungi, a situation quite similar to the one found after biomass pre-treatments in biofuel production process.