Riassunto analitico
Lytic Polysaccharide Monooxygenases (LPMOs) , are a class of mono-copper metalloenzymes identified in 1992, that can depolymerize recalcitrant polysaccharide matrices. The main effect they caused is the hydroxylation at the C1 or C4 carbon atom, followed by cleavage of the glycosidic bond, resulting in the formation of oxidized oligomers or monomers of the constituent sugars, thus exposing more chain ends as 'access points' to glycosidic hydrolyse (GH), resulting in increased saccharification. They all have a catalytic mono-copper catalytic centre and a β-sandwich fold, similar to immunoglobulins within the protein core. This thesis project focuses on the production, purification and characterisation of two Lytic Polysaccharide Monooxygenases, ScLPMOAA10B and ScLPMOAA10C. These enzymes are crucial for breaking down sugar-based biopolymers such as cellulose and chitin, making them highly valuable for biomass conversion. Furthermore, by exploiting the oxidation of the C-O-C bond at the C1 or C4 position of a sugar chain, these enzymes can be employed in developing enzyme cocktails for biomass degradation. We have produced ScLPMOAA10B and ScLPMOAA10C in E. coli, optimized a protocol for their purification and confirmed their activity by using ATR-FTIR spectroscopy and ESI-Ion Trap mass spectrometry.
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Abstract
Lytic Polysaccharide Monooxygenases (LPMOs) , are a class of mono-copper metalloenzymes identified in 1992, that can depolymerize recalcitrant polysaccharide matrices. The main effect they caused is the hydroxylation at the C1 or C4 carbon atom, followed by cleavage of the glycosidic bond, resulting in the formation of oxidized oligomers or monomers of the constituent sugars, thus exposing more chain ends as 'access points' to glycosidic hydrolyse (GH), resulting in increased saccharification. They all have a catalytic mono-copper catalytic centre and a β-sandwich fold, similar to immunoglobulins within the protein core. This thesis project focuses on the production, purification and characterisation of two Lytic Polysaccharide Monooxygenases, ScLPMOAA10B and ScLPMOAA10C. These enzymes are crucial for breaking down sugar-based biopolymers such as cellulose and chitin, making them highly valuable for biomass conversion. Furthermore, by exploiting the oxidation of the C-O-C bond at the C1 or C4 position of a sugar chain, these enzymes can be employed in developing enzyme cocktails for biomass degradation. We have produced ScLPMOAA10B and ScLPMOAA10C in E. coli, optimized a protocol for their purification and confirmed their activity by using ATR-FTIR spectroscopy and ESI-Ion Trap mass spectrometry.
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