Mosbech C

References (3)

Title : The structural basis of fungal glucuronoyl esterase activity on natural substrates - Ernst_2020_Nat.Commun_11_1026
Author(s) : Ernst HA , Mosbech C , Langkilde AE , Westh P , Meyer AS , Agger JW , Larsen S
Ref : Nat Commun , 11 :1026 , 2020
Abstract : Structural and functional studies were conducted of the glucuronoyl esterase (GE) from Cerrena unicolor (CuGE), an enzyme catalyzing cleavage of lignin-carbohydrate ester bonds. CuGE is an alpha/beta-hydrolase belonging to carbohydrate esterase family 15 (CE15). The enzyme is modular, comprised of a catalytic and a carbohydrate-binding domain. SAXS data show CuGE as an elongated rigid molecule where the two domains are connected by a rigid linker. Detailed structural information of the catalytic domain in its apo- and inactivated form and complexes with aldouronic acids reveal well-defined binding of the 4-O-methyl-a-D-glucuronoyl moiety, not influenced by the nature of the attached xylo-oligosaccharide. Structural and sequence comparisons within CE15 enzymes reveal two distinct structural subgroups. CuGE belongs to the group of fungal CE15-B enzymes with an open and flat substrate-binding site. The interactions between CuGE and its natural substrates are explained and rationalized by the structural results, microscale thermophoresis and isothermal calorimetry.
ESTHER : Ernst_2020_Nat.Commun_11_1026
PubMedSearch : Ernst_2020_Nat.Commun_11_1026
PubMedID: 32094331
Gene_locus related to this paper: cerui-gce

Title : Enzyme kinetics of fungal glucuronoyl esterases on natural lignin-carbohydrate complexes - Mosbech_2019_Appl.Microbiol.Biotechnol_103_4065
Author(s) : Mosbech C , Holck J , Meyer A , Agger JW
Ref : Applied Microbiology & Biotechnology , 103 :4065 , 2019
Abstract : Glucuronoyl esterases (CE15 family) enable targeted cleavage of ester linkages in lignin-carbohydrate complexes (LCCs), particularly those linking lignin and glucuronoyl residues in xylan. A substantial challenge in characterization and kinetic analysis of CE15 enzymes has been the lack of proper substrates. Here, we present an assay using an insoluble LCC-rich lignin fraction from birch; lignin-rich pellet (LRP). The assay employs quantification of enzyme reaction products by LC-MS. The kinetics of four fungal CE15 enzymes, PsGE, CuGE, TtGE, and AfuGE originating from lignocellulose-degrading fungi Punctularia strigosozonata, Cerrena unicolor, Thielavia terrestris, and Armillaria fuscipes respectively were characterized and compared using this new assay. All four enzymes had activity on LRP and showed a clear preference for the insoluble substrate compared with smaller soluble LCC mimicking esters. End-product profiles were near identical for the four enzymes but differences in kinetic parameters were observed. TtGE possesses an alternative active site compared with the three other enzymes as it has the position of the catalytic glutamic acid occupied by a serine. TtGE performed poorly compared with the other enzymes. We speculate that glucuronoyl LCCs are not the preferred substrate of TtGE. Removal of an N-terminal CBM on CuGE affected the catalytic efficiently of the enzyme by reducing K(cat) by more than 30%. Reaction products were detected from all four CE15s on a similar substrate from spruce indicating a more generic GE activity not limited to the hardwood. The assay with natural substrate represents a novel tool to study the natural function and kinetics of CE15s.
ESTHER : Mosbech_2019_Appl.Microbiol.Biotechnol_103_4065
PubMedSearch : Mosbech_2019_Appl.Microbiol.Biotechnol_103_4065
PubMedID: 30949809

Title : The natural catalytic function of CuGE glucuronoyl esterase in hydrolysis of genuine lignin-carbohydrate complexes from birch - Mosbech_2018_Biotechnol.Biofuels_11_71
Author(s) : Mosbech C , Holck J , Meyer AS , Agger JW
Ref : Biotechnol Biofuels , 11 :71 , 2018
Abstract : BACKGROUND: Glucuronoyl esterases belong to carbohydrate esterase family 15 and catalyze de-esterification. Their natural function is presumed to be cleavage of ester linkages in lignin-carbohydrate complexes particularly those linking lignin and glucuronoyl residues in xylans in hardwood. RESULTS: Here, we show for the first time a detailed product profile of aldouronic acids released from birchwood lignin by a glucuronoyl esterase from the white-rot fungus Cerrena unicolor (CuGE). CuGE releases substrate for GH10 endo-xylanase which results in significantly increased product release compared to the action of endo-xylanase alone. CuGE also releases neutral xylo-oligosaccharides that can be ascribed to the enzymes feruloyl esterase side activity as demonstrated by release of ferulic acid from insoluble wheat arabinoxylan. CONCLUSION: The data verify the enzyme's unique ability to catalyze removal of all glucuronoxylan associated with lignin and we propose that this is a direct result of enzymatic cleavage of the ester bonds connecting glucuronoxylan to lignin via 4-O-methyl glucuronoyl-ester linkages. This function appears important for the fungal organism's ability to effectively utilize all available carbohydrates in lignocellulosic substrates. In bioprocess perspectives, this enzyme is a clear candidate for polishing lignin for residual carbohydrates to achieve pure, native lignin fractions after minimal pretreatment.
ESTHER : Mosbech_2018_Biotechnol.Biofuels_11_71
PubMedSearch : Mosbech_2018_Biotechnol.Biofuels_11_71
PubMedID: 29560026
Gene_locus related to this paper: cerui-gce