In situ imaging of LPMO action on plant tissues.

Lytic polysaccharide monooxygenases (LPMOs) are copper-dependent enzymes that oxidatively cleave recalcitrant polysaccharides such as cellulose. Several studies have reported LPMO action in synergy with other carbohydrate-active enzymes (CAZymes) for the degradation of lignocellulosic biomass but direct LPMO action at the plant tissue level remains challenging to investigate. Here, we have developed a MALDI-MS imaging workflow to detect oxidised oligosaccharides released by a cellulose-active LPMO at cellular level on maize tissues.

The disordered C-terminal tail of fungal LPMOs from phytopathogens mediates protein dimerization and impacts plant penetration.

Lytic polysaccharide monooxygenases (LPMOs) are monocopper enzymes that oxidatively degrade various polysaccharides, such as cellulose. Despite extensive research on this class of enzymes, the role played by their C-terminal regions predicted to be intrinsically disordered (dCTR) has been overlooked. Here, we investigated the function of the dCTR of an LPMO, called AA9A, up-regulated during plant infection by , the causative agent of anthracnose.

Methionine oxidation of carbohydrate-active enzymes during white-rot wood decay.

White-rot fungi employ secreted carbohydrate-active enzymes (CAZymes) along with reactive oxygen species (ROS), like hydrogen peroxide (HO), to degrade lignocellulose in wood. HO serves as a co-substrate for key oxidoreductases during the initial decay phase. While the degradation of lignocellulose by CAZymes is well documented, the impact of ROS on the oxidation of the secreted proteins remains unclear, and the identity of the oxidized proteins is unknown.

Insights into peculiar fungal LPMO family members holding a short C-terminal sequence reminiscent of phosphate binding motifs.

Lytic polysaccharide monooxygenases (LPMOs) are taxonomically widespread copper-enzymes boosting biopolymers conversion (e.g. cellulose, chitin) in Nature.

Revisiting the AA14 family of lytic polysaccharide monooxygenases and their catalytic activity.

Lytic polysaccharide monooxygenases (LPMOs) belonging to the AA14 family are believed to contribute to the enzymatic degradation of lignocellulosic biomass by specifically acting on xylan in recalcitrant cellulose-xylan complexes. Functional characterization of an AA14 LPMO from Trichoderma reesei, TrAA14A, and a re-evaluation of the properties of the previously described AA14 from Pycnoporus coccineus, PcoAA14A, showed that these proteins have oxidase and peroxidase activities that are common for LPMOs. However, we were not able to detect activity on cellulose-associated xylan or any other tested polysaccharide substrate, meaning that the substrate of these enzymes remains unknown.

Optimized Lytic Polysaccharide Monooxygenase Action Increases Fiber Accessibility and Fibrillation by Releasing Tension Stress in Cellulose Cotton Fibers.

Lytic polysaccharide monooxygenase (LPMO) enzymes have recently shaken up our knowledge of the enzymatic degradation of biopolymers and cellulose in particular. This unique class of metalloenzymes cleaves cellulose and other recalcitrant polysaccharides using an oxidative mechanism. Despite their potential in biomass saccharification and cellulose fibrillation, the detailed mode of action of LPMOs at the surface of cellulose fibers still remains poorly understood and highly challenging to investigate.

A fungal lytic polysaccharide monooxygenase is required for cell wall integrity, thermotolerance, and virulence of the fungal human pathogen Cryptococcus neoformans.

Fungi often adapt to environmental stress by altering their size, shape, or rate of cell division. These morphological changes require reorganization of the cell wall, a structural feature external to the cell membrane composed of highly interconnected polysaccharides and glycoproteins. Lytic polysaccharide monooxygenases (LPMOs) are copper-dependent enzymes that are typically secreted into the extracellular space to catalyze initial oxidative steps in the degradation of complex biopolymers such as chitin and cellulose.

A special issue of Essays in Biochemistry on current advances about CAZymes and their impact and key role in human health and environment.

Carbohydrate active enzymes (CAZymes) and their biochemical characterization have been the subject of extensive research over the past ten years due to their importance to carbohydrate metabolism in different biological contexts. For instance, the understanding that 'polysaccharide utilizing loci' (PUL) systems hosted by specific 'carbohydrate degraders' in the intestinal microbiota play key roles in health and disease, such as Crohn's disease, ulcerative colitis or colorectal cancer to name the most well-characterized, has led to an outstanding effort in trying to decipher the molecular mechanisms by which these processes are organized and regulated. The past 10 years has also seen the expansion of CAZymes with auxiliary activities, such as lytic polysaccharide monooxygenases (LPMOs) or even sulfatases, and interest has grown in general about the enzymes needed to remove the numerous decorations and modifications of complex biomass, such as carbohydrate esterases (CE).

Carbohydrate-active enzymes in animal feed.

Considering an ever-growing global population, which hit 8 billion people in the fall of 2022, it is essential to find solutions to avoid the competition between human food and animal feed for croplands. Agricultural co-products have become important components of the circular economy with their use in animal feed. Their implementation was made possible by the addition of exogenous enzymes in the diet, especially carbohydrate-active enzymes (CAZymes).

Rational engineering of AA5_2 copper radical oxidases to probe the molecular determinants governing their substrate selectivity.

Fungal copper radical oxidases (CROs) from the Auxiliary Activity family 5 (AA5) constitute a group of metalloenzymes that oxidize a wide panel of natural compounds, such as galactose-containing saccharides or primary alcohols, into product derivatives exhibiting promising biotechnological interests. Despite a well-conserved first copper-coordination sphere and overall fold, some members of the AA5_2 subfamily are incapable of oxidizing galactose and galactosides but conversely efficiently catalyse the oxidation of diverse aliphatic alcohols. The objective of this study was to understand which residues dictate the substrate preferences between alcohol oxidases and galactose oxidases within the AA5_2 subfamily.

Tandem metalloenzymes gate plant cell entry by pathogenic fungi.

Global food security is endangered by fungal phytopathogens causing devastating crop production losses. Many of these pathogens use specialized appressoria cells to puncture plant cuticles. Here, we unveil a pair of alcohol oxidase-peroxidase enzymes to be essential for pathogenicity.

The Maize Pathogen Ustilago maydis Secretes Glycoside Hydrolases and Carbohydrate Oxidases Directed toward Components of the Fungal Cell Wall.

Filamentous fungi are keystone microorganisms in the regulation of many processes occurring on Earth, such as plant biomass decay and pathogenesis as well as symbiotic associations. In many of these processes, fungi secrete carbohydrate-active enzymes (CAZymes) to modify and/or degrade carbohydrates. Ten years ago, while evaluating the potential of a secretome from the maize pathogen Ustilago maydis to supplement lignocellulolytic cocktails, we noticed it contained many unknown or poorly characterized CAZymes.

A simple and direct ionic chromatography method to monitor galactose oxidase activity.

Galactose oxidase (GalOx, EC.1.1.

2D and 3D maximum-quantum NMR and diffusion spectroscopy for the characterization of enzymatic reaction mixtures.

1D H NMR spectroscopy has been widely used to monitor enzymatic activity by recording the evolution of the spectra of substrates and/or products, thanks to the linear response of NMR. For complex systems involving the coexistence of multiple compounds (substrate, final product and various intermediates), the identification and quantification can be a more arduous task. Here, we present a simple analytical method for the rapid characterization of reaction mixtures involving enzymatic complexes using Maximum Quantum (MaxQ) NMR, accelerated with the Non-Uniform Sampling (NUS) acquisition procedure.

Predicting Protein Conformational Disorder and Disordered Binding Sites.

In the last two decades it has become increasingly evident that a large number of proteins adopt either a fully or a partially disordered conformation. Intrinsically disordered proteins are ubiquitous proteins that fulfill essential biological functions while lacking a stable 3D structure. Their conformational heterogeneity is encoded by the amino acid sequence, thereby allowing intrinsically disordered proteins or regions to be recognized based on their sequence properties.

Activity-based protein profiling reveals dynamic substrate-specific cellulase secretion by saprotrophic basidiomycetes.

Background: Fungal saccharification of lignocellulosic biomass occurs concurrently with the secretion of a diverse collection of proteins, together functioning as a catalytic system to liberate soluble sugars from insoluble composite biomaterials. How different fungi respond to different substrates is of fundamental interest to the developing biomass saccharification industry. Among the cornerstones of fungal enzyme systems are the highly expressed cellulases (endo-β-glucanases and cellobiohydrolases).

Tunable Production of ()- or ()-Citronellal from Geraniol via a Bienzymatic Cascade Using a Copper Radical Alcohol Oxidase and Old Yellow Enzyme.

Biocatalytic pathways for the synthesis of (-)-menthol, the most sold flavor worldwide, are highly sought-after. To access the key intermediate ()-citronellal used in current major industrial production routes, we established a one-pot bienzymatic cascade from inexpensive geraniol, overcoming the problematic biocatalytic reduction of the mixture of ()-isomers in citral by harnessing a copper radical oxidase (AlcOx) and an old yellow enzyme (OYE). The cascade using OYE2 delivered 95.

The ectomycorrhizal basidiomycete Laccaria bicolor releases a GH28 polygalacturonase that plays a key role in symbiosis establishment.

In ectomycorrhiza, root penetration and colonization of the intercellular space by symbiotic hyphae is thought to rely on the mechanical force that results from hyphal tip growth, enhanced by the activity of secreted cell-wall-degrading enzymes. Here, we characterize the biochemical properties of the symbiosis-induced polygalacturonase LbGH28A from the ectomycorrhizal fungus Laccaria bicolor. The transcriptional regulation of LbGH28A was measured by quantitative PCR (qPCR).

On the expansion of biological functions of lytic polysaccharide monooxygenases.

Lytic polysaccharide monooxygenases (LPMOs) constitute an enigmatic class of enzymes, the discovery of which has opened up a new arena of riveting research. LPMOs can oxidatively cleave the glycosidic bonds found in carbohydrate polymers enabling the depolymerisation of recalcitrant biomasses, such as cellulose or chitin. While most studies have so far mainly explored the role of LPMOs in a (plant) biomass conversion context, alternative roles and paradigms begin to emerge.