A5005199010- Electrochemical sensors and biosensors
- Enzyme-mediated dye degradation
- Electrochemical Analysis and Applications
- Photosynthetic Processes and Mechanisms
- Biofuel production and bioconversion
- Enzyme Catalysis and Immobilization
- Microbial Metabolic Engineering and Bioproduction
- Enzyme Production and Characterization
- Microbial Fuel Cells and Bioremediation
- Microbial Metabolism and Applications
- Microbial bioremediation and biosurfactants
- Hemoglobin structure and function
- Protein Structure and Dynamics
- Lignin and Wood Chemistry
- Metal-Catalyzed Oxygenation Mechanisms
- Advanced biosensing and bioanalysis techniques
- Porphyrin Metabolism and Disorders
- Heme Oxygenase-1 and Carbon Monoxide
- Polysaccharides and Plant Cell Walls
- Fungal and yeast genetics research
- Biochemical and biochemical processes
- Microfluidic and Capillary Electrophoresis Applications
BOKU University
2012-2023
Ninety percent of lignocellulose-degrading fungi contain genes encoding lytic polysaccharide monooxygenases (LPMOs). These enzymes catalyze the initial oxidative cleavage recalcitrant polysaccharides after activation by an electron donor. Understanding source electrons is fundamental to fungal physiology and will also help with exploitation LPMOs for biomass processing. Using genome data biochemical methods, we characterized compared different extracellular sources LPMOs: cellobiose...
ABSTRACT The genome of Neurospora crassa encodes two different cellobiose dehydrogenases (CDHs) with a sequence identity only 53%. So far, CDH IIA, which is induced during growth on cellulose and features C-terminal carbohydrate binding module (CBM), was detected in the secretome N. preliminarily characterized. IIB not significantly upregulated cellulosic material lacks CBM. Since could be identified secretome, both CDHs were recombinantly produced Pichia pastoris . With cytochrome...
Abstract Lytic polysaccharide monooxygenase (LPMO) supports biomass hydrolysis by increasing saccharification efficiency and rate. Recent studies demonstrate that H 2 O rather than is the cosubstrate of LPMO-catalyzed depolymerization polysaccharides. Some have questioned physiological relevance -based mechanism for plant cell wall degradation. This study reports localized time-resolved determination LPMO activity on poplar wood walls measuring concentration in their vicinity with a...
Abstract Lytic polysaccharide monooxygenases (LPMOs) represent a recent addition to the carbohydrate‐active enzymes and are classified as auxiliary activity (AA) families 9, 10, 11, 13. LPMOs crucial for effective degradation of recalcitrant polysaccharides like cellulose or chitin. These copper‐dependent utilize redox mechanism cleave glycosidic bonds that is dependent on molecular oxygen an external electron donor. The electrons can be provided by various sources, such chemical compounds...
In past years, new lytic polysaccharide monooxygenases (LPMOs) have been discovered as distinct in their substrate specificity. Their unconventional, surface-exposed catalytic sites determine enzymatic activities, while binding govern recognition and regioselectivity. An additional factor influencing activity is the presence or absence of a family 1 carbohydrate module (CBM1) connected via linker to C-terminus LPMO. This study investigates changes induced by shortening second active site...
Lytic polysaccharide monooxygenases (LPMOs) are powerful enzymes that oxidatively cleave plant cell wall polysaccharides. LPMOs classified as fungal Auxiliary Activities family 9 (AA9) have been mainly studied for their activity towards cellulose; however, various members of this AA9 also shown to hemicelluloses, in particularly xyloglucan (XG). So far, it has not detail how act XG degradation, and particular, the mode-of-action relates structural configuration these LPMOs. Two Neurospora...
Fungal high redox potential laccases are proposed as cathodic biocatalysts in implantable enzymatic fuel cells to generate cell voltages. Their application is limited mainly through their acidic pH optimum and chloride inhibition. This work investigates evolutionary engineering strategies increase the of a chloride-tolerant, laccase from ascomycete Botrytis aclada. The was subjected two rounds directed evolution clones screened for increased stability activity at 6.5. Beneficial mutation...
The natural function of cellobiose dehydrogenase (CDH) to donate electrons from its catalytic flavodehydrogenase (DH) domain via cytochrome (CYT) lytic polysaccharide monooxygenase (LPMO) is an example a highly efficient extracellular electron transfer chain. To investigate the CYT movement in two occurring steps, CDHs ascomycete Neurospora crassa (NcCDHIIA and NcCDHIIB) five chimeric CDH enzymes created by swapping were studied combination with fungus' own LPMOs (NcLPMO9C NcLPMO9F). Kinetic...
Enzymatic hydrolysis of lignocellulosic biomass for biofuel production relies on complex multi-enzyme ensembles. Continuous and accurate measurement the released key products is crucial in optimizing industrial degradation process also investigating activity interaction between involved enzymes insoluble substrate. Amperometric biosensors have been applied to perform continuous cellobiose measurements during enzymatic pure cellulose powders. The oxygen-sensitive mediators used these...
Recently, a novel pathway for heme b biosynthesis in Gram-positive bacteria has been proposed. The final poorly understood step is catalyzed by an enzyme called HemQ and includes two decarboxylation reactions leading from coproheme to b. Coproheme suggested act as both substrate redox active cofactor this reaction. In the study presented here, we focus on HemQs Listeria monocytogenes (LmHemQ) Staphylococcus aureus (SaHemQ) recombinantly produced apoproteins Escherichia coli. We demonstrate...
Large-scale protein domain dynamics and electron transfer are often associated. However, as motions span a broad range of time length scales, it is challenging to identify thus link functionally relevant dynamic changes in proteins. It hypothesized that large-scale direct electrons through FAD heme b cofactor the fungal cellobiose dehydrogenase (CDH) enzymes type-II copper center (T2Cu) polysaccharide-degrading lytic polysaccharide monooxygenases (LPMOs). yet, CDH have not been linked...
Cytochromes are a large and well-known group of electron transfer proteins, but only very few used in bioelectrochemistry, e.g. to serve as shuttles between biocatalysts electrodes. There, cytochromes generate an artificial pathway if their structural electrochemical properties fitting. considered suitable small feature exposed heme cofactor, redox potential, electrochemically reversible process, sufficient protein production, fast other enzymes hemoproteins. Out 850 screened hemoproteins,...
Glyoxal oxidase (GLOX) is an extracellular source of H2O2 in white-rot secretomes, where it acts concert with peroxidases to degrade lignin. It has been reported that GLOX requires activation prior catalytic turnover and a peroxidase system can fulfill this task. In study, we verify oxidation product horseradish peroxidase, the radical cation 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), activator for GLOX. A spectroelectrochemical cell was used generate activating species,...
The development of third generation biosensors depends on the availability direct electron transfer (DET) capable enzymes. A successful strategy is to fuse a cytochrome domain an enzyme fulfil function built-in redox mediator between catalytic center and electrode. In this study, we fused Neurospora crassa CDH IIA (NcCYT) N-terminally glucose dehydrogenase from Glomerella cingulata (GcGDH) generate chimeric NcCYT-GcGDH in large amount for further studies. Heterologous expression P. pastoris...
Abstract Cellobiose dehydrogenase (CDH) is a bioelectrocatalyst that enables direct electron transfer (DET) in biosensors and biofuel cells. The application of this bidomain hemoflavoenzyme for physiological glucose measurements limited by its acidic pH optimum slow interdomain (IET) at 7.5. reason rate‐limiting step electrostatic repulsion the interface between catalytic domain mediating cytochrome (CYT). We applied rational engineering to accelerate IET prevailing blood or interstitial...
Abstract The function of cellobiose dehydrogenase (CDH) in biosensors, biofuel cells, and as a physiological redox partner lytic polysaccharide monooxygenase (LPMO) is based on its role an electron donor. Before donating electrons to LPMO or electrodes, interdomain transfer from the catalytic FAD‐containing domain shuttling cytochrome CDH required. This study investigates two crucial amino acids located at interaction by structure‐based engineering. kinetics wild‐type Myriococcum...
Current processes for lignocellulose deconstruction are unspecific and produce some constituents in poor quality. Specific biocatalysts could achieve optimal segregation together with minimal damage to cellulose lignin provide high-quality feedstocks industry. Naturally occurring fungal oxidoreductases perform this task, but their characterisation - hence optimisation industrial application is difficult because of the experimental challenges. The mission OXIDISE develop appropriate methods...