Polysaccharide Degrading Biocatalysts

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The transformation of polysaccharides into valuable compounds for health and industry requires the careful application of enzyme protocols and controlled biocatalysis.

Polysaccharide-Degrading Biocatalysts provides a thorough grounding in these biocatalytic processes and their growing role in the depolymerization of polysaccharides, empowering researchers to discover and develop new enzyme-based approaches across pharmaceuticals, fuels, and food engineering. Here, over a dozen leading experts offer a close examination of structural polysaccharides, genetic modification of polysaccharides, polysaccharide degradation routes, pretreatments for enzymatic hydrolysis, hemicellulose-degrading enzymes, biomass valorization processes, oligosaccharide production, and enzyme immobilization for the hydrolysis of polysaccharides, among other topics and related research protocols. A final chapter considers perspectives and challenges in an evolving, carbohydrate-based economy.

Author(s): Rosana Goldbeck, Patricia Poletto
Series: Foundations and Frontiers in Enzymology
Publisher: Academic Press
Year: 2023

Language: English
Pages: 452
City: London

Front Cover
Polysaccharide-Degrading Biocatalysts
Copyright
Contents
Contributors
Chapter 1: Plant cell wall polysaccharides: Methodologies for compositional, structural, and physicochemical characterization
1. Introduction to the analysis of plant cell wall polysaccharides
2. Sample preparation for the polysaccharides analysis
3. Chemical analysis of plant cell wall polysaccharides-Glycosyl residues composition
3.1. High-performance liquid chromatography (HPLC)
3.2. High-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD)
3.3. Gas chromatography-The alditol acetate (AA) and trimethylsilyl (TMS) derivatives
4. Structural analysis of plant cell wall polysaccharides
4.1. Nuclear magnetic resonance spectroscopy (NMR)
4.2. Gas chromatography-Methylation analysis
4.3. Immunological approaches for structural polysaccharides characterization
5. Complementary methods for the analysis of plant cell wall polysaccharides
5.1. Atomic force microscopy (AFM)
5.2. Electron microscopy
5.3. X-ray diffraction (XRD) analysis
5.4. Fourier-transform infrared spectroscopy (FTIR)
6. Concluding remarks
References
Chapter 2: Genetic modification of plants to increase the saccharification of lignocellulose
1. Introduction
2. Lignin biosynthesis
3. Molecular approaches to cell wall modification
3.1. Genetic modification in cellulose biosynthesis
3.2. Genetic modification in hemicellulose biosynthesis
3.3. Genetic modification in lignin biosynthesis
4. Technologies for genetic modification of the cell wall
4.1. Antisense oligonucleotide
4.2. RNAi
4.3. Transcription activator-like effector nucleases (TALEN)
4.4. CRISPR/Cas9
5. Final considerations
References
Chapter 3: The diversity of plant carbohydrate hydrolysis in nature and technology
1. Introduction
2. Types of hydrolysis
2.1. Acid hydrolysis
2.2. Enzymatic hydrolysis
3. Plant sugars
3.1. Sucrose, raffinose, and fructans
3.2. Starch
3.3. Cell wall polysaccharides
3.3.1. Exogenous degradation of cell walls: Hydrolysis by microorganisms and animals
3.3.2. Endogenous degradation of cell walls
3.3.2.1. Abscission
3.3.2.2. Fruit ripening
3.3.2.3. Aerenchyma
3.3.2.4. Long-term storage heteropolysaccharides mobilization
4. Concluding remarks
Acknowledgments
References
Chapter 4: State-of-the-art experimental and computational approaches to investigate structure, substrate recognition, an ...
1. Sample preparation for structural and biophysical analyses
1.1. Molecular cloning strategies for enzyme expression
1.2. Enzyme purification for chemical and structural homogeneity
2. Methods to analyze enzyme stability and structural homogeneity
2.1. Dynamic light scattering
2.2. Circular dichroism spectroscopy
2.3. Intrinsic and extrinsic fluorescence
3. Methods to analyze protein conformation and oligomerization
3.1. Size-exclusion chromatography with multiangle light scattering (SEC-MALS)
3.2. Analytical ultracentrifugation
3.3. Small-angle X-ray scattering
4. Methods to analyze enzyme-substrate interactions
4.1. Isothermal titration calorimetry (ITC)
4.2. Microscale thermophoresis (MST)
4.3. Chemical cross-linking mass spectrometry (XL-MS)
4.4. Hydrogen-deuterium exchange mass spectrometry (HDX-MS)
5. Experimental approaches for structure elucidation
5.1. X-ray crystallography (XTAL)
5.1.1. Crystallization
5.1.2. X-ray source, data collection, and structure determination
5.1.3. Room temperature and time-resolved serial crystallography
5.2. Single-particle cryo-EM at atomic resolution
5.3. NMR spectroscopy
6. In silico methods for structural analysis
6.1. Artificial intelligence-based modeling (Alpha-fold/RosettaFold)
6.2. Molecular docking: Modeling of enzyme-ligand binding
6.3. QM/MM-based simulations to analyze biocatalytic reactions
References
Chapter 5: Pretreatments as a key for enzymatic hydrolysis of lignocellulosic biomass
1. Introduction
2. Factors affecting enzymatic hydrolysis and their relationship with the pretreatment
2.1. Biomass physical and chemical factors
2.1.1. Accessible, specific, and internal surface area
2.1.2. Cellulose crystallinity and degree of polymerization
2.1.3. Chemical composition
2.1.3.1. Pretreatments promoting hemicellulose removal
2.1.3.2. Delignified pretreated biomass
2.1.3.3. Effect of residual lignin on enzymatic hydrolysis
2.2. Enzyme inhibitors produced in the pretreatment step
3. Pretreatment of lignocellulosic biomass
3.1. Physical pretreatments
3.1.1. Mechanical pretreatment
3.1.2. Ultrasound pretreatment
3.1.3. Microwave pretreatment
3.2. Chemical and physicochemical pretreatments
3.2.1. Acid pretreatment
3.2.2. Hydrothermal and steam explosion pretreatments
3.2.3. Alkaline pretreatment
3.2.4. Organosolv pretreatment
3.2.5. Ionic liquids
3.2.6. Eutectic solvents
3.3. Biological pretreatments
4. Conclusions
References
Chapter 6: Importance of accessory enzymes in hemicellulose degradation
1. Introduction
2. Xylanolytic enzymes
2.1. Enzymes acting on the side chains
2.1.1. α-Glucuronidase
2.1.1.1. Reaction catalyzed and classification
2.1.1.2. Substrate specificity
2.1.2. α-l-Arabinofuranosidase
2.1.2.1. Reaction catalyzed
2.1.2.2. Classification
2.1.2.3. Substrate specificity and reaction mechanism
2.1.3. Carbohydrate esterases
2.1.3.1. Acetylxylan esterase
2.1.3.1.1. Reaction catalyzed
2.1.3.1.2. Classification
2.1.3.1.3. Substrate specificity and reaction mechanism
2.1.3.2. Feruloyl esterase
2.1.3.2.1. Reaction catalyzed
2.1.3.2.2. Classification, substrate specificity, and reaction mechanism
2.1.3.3. Glucuronoyl esterase
2.1.3.3.1. Reaction catalyzed
2.1.3.3.2. Classification, structure, and reaction mechanism
2.1.4. Rare debranching enzymes
2.1.4.1. α-Xylosidase
2.1.4.1.1. Reaction catalyzed
2.1.4.1.2. Classification, substrate specificity, and reaction mechanism
2.1.4.2. α-l-Galactosidase
2.1.4.2.1. Reaction catalyzed
2.1.4.2.2. Classification, substrate specificity, and reaction mechanism
2.1.4.3. β-1,3-Xylosidase
2.1.4.3.1. Reaction catalyzed
2.2. Exo-acting xylanolytic enzymes attacking the main chain
2.2.1. β-Xylosidase
2.2.1.1. Reaction catalyzed
2.2.1.2. Classification and reaction mechanism
2.2.1.3. Substrate specificity
2.2.2. Exo-β-1,4-xylanase
2.2.3. Xylobiohydrolase
2.2.3.1. Reaction catalyzed
2.2.3.2. Classification, substrate specificity, and reaction mechanism
2.2.4. Reducing-end xylose-releasing enzyme (Rex)
2.2.4.1. Reaction catalyzed
2.2.4.2. Classification, substrate specificity, and reaction mechanism
3. Mannanolytic enzymes
3.1. Enzymes acting on the side chains
3.1.1. α-Galactosidase
3.1.1.1. Reaction catalyzed
3.1.1.2. Classification
3.1.1.3. Substrate specificity and reaction mechanism
3.1.2. Acetylmannan esterase
3.1.2.1. Reaction catalyzed
3.1.2.2. Classification
3.1.2.3. Substrate specificity and reaction mechanism
3.2. Exo-acting mannanolytic enzymes attacking the main chain
3.2.1. β-Mannosidase
3.2.1.1. Reaction catalyzed
3.2.1.2. Classification, substrate specificity, and reaction mechanism
3.2.2. Mannobiohydrolase
3.2.2.1. Reaction catalyzed
3.2.2.2. Classification, substrate specificity, and reaction mechanism
3.2.3. β-Glucosidase
3.2.3.1. Reaction catalyzed
3.2.3.2. Classification, substrate specificity, and reaction mechanism
3.2.4. Phosphorylases acting on β-manno-based substrates
3.2.4.1. Mannosylglucose phosphorylases
3.2.4.1.1. Reaction catalyzed
3.2.4.1.2. Classification
3.2.4.1.3. Substrate specificity and reaction mechanism
3.2.4.2. β-1,4-Mannooligosaccharide phosphorylases
3.2.4.2.1. Reaction catalyzed
3.2.4.2.2. Classification
3.2.4.2.3. Substrate specificity and reaction mechanism
4. Conclusions
Funding
References
Chapter 7: How ligninolytic enzymes can help in the degradation of biomass polysaccharides, cleavage, and catalytic mecha ...
1. Lignocellulosic biomass
2. Lignin
3. Ligninases
3.1. Laccases (Lacs)
3.2. Lignin peroxidase (LiP)
3.3. Manganese peroxidases (MnPs)
3.4. Versatile peroxidase (VP)
4. Ligninolytic enzymes and prospects
References
Chapter 8: Biochemical and biotechnological aspects of microbial amylases
1. Introduction
2. Amylase: The starch-digesting enzyme
2.1. α-Amylase: Structure and mechanism of action
2.2. β-Amylase
2.3. γ-Amylase (EC 3.2.1.3)
3. Commercial production of α-amylases
4. Applications of α-amylase
4.1. Food industry
4.2. Textile industry
4.3. Bio-fuel production
4.4. Detergent industry
4.5. Paper industry
4.6. Other promising applications
4.7. Biomedical significances
5. Conclusion and future perspectives
References
Chapter 9: Hydrolysis of complex pectin structures: Biocatalysis and bioproducts
1. Introduction
2. Pectin complex structure
2.1. Homogalacturonan (HG)
2.2. Xylogalacturonans
2.3. Rhamnogalacturonan-I (RG-I)
2.4. Rhamnogalacturonan-II (RG-II)
3. Types of pectins
4. Sources of pectin
5. Pectin: Diverse uses
6. Pectinases
6.1. Protopectinase
6.2. Esterase
6.3. Depolymerases
6.4. Polygalacturonase
6.5. Lyases
7. Structural aspects of protein families related to pectin degradation
8. Conclusion
Acknowledgment
References
Chapter 10: Macroalgal polysaccharides: Biocatalysts in biofuel/bioenergy production
1. Introduction
2. The bio-refinery concept
3. Algae and its classification
3.1. Microalgae
3.2. Macroalgae
3.2.1. Green algal polysaccharides
3.2.2. Red algal polysaccharides
3.2.2.1. Cellulose
3.2.2.2. Mannan
3.2.2.3. Xylan
3.2.2.4. Sulfated galactans
3.2.2.4.1. Agar
3.2.2.4.2. Carrageenan
3.2.3. Brown algal polysaccharides
3.2.3.1. Alginate
3.2.3.2. Fucoidan
3.2.3.3. Laminarin
3.2.3.4. Mannitol
4. Extraction of macroalgal polysaccharides
4.1. Mechanical treatment
4.1.1. Size reduction
4.1.2. Beating
4.1.3. Washing
4.1.4. Ultrasound-assisted extraction (UAE)
4.2. Thermal treatment
4.2.1. Microwave
4.2.2. Steam explosion
4.2.3. Pressurized liquid extraction (PLE)
4.2.4. Other thermal pre-treatments
4.3. Chemical treatment
4.3.1. Alkali or acid treatment
4.3.2. Peroxide treatment
4.4. Biological treatment
5. Biocatalysts in bio-refinery and biofuel production
5.1. Bioethanol production
5.1.1. Pre-treatment
5.1.2. Hydrolysis
5.1.3. Fermentation
5.1.4. Recovery processes
5.1.5. Researches related to the use of enzymes in ethanol production from different macroalgae
5.2. Biobutanol production
5.3. Biogas production
6. Conclusions and future prospects
Acknowledgments
References
Chapter 11: Mathematical modeling of the enzymatic hydrolysis of polysaccharides: A primer
1. Aims and scope of this chapter
2. Scales at which the enzymatic hydrolysis of polysaccharides can be modeled
2.1. Atomic-scale models of interaction between enzymes and polysaccharides
2.2. Molecular-scale models of interaction between enzymes and polysaccharides
2.3. Dynamic macroscale models of enzymatic hydrolysis processes
2.4. Models used specifically as tools in parameter estimation
3. Features of substrates, enzymes, and models
3.1. Features of polysaccharide substrates
3.2. Features of enzymes used to hydrolyze polymeric substrates
3.3. Types of models
3.3.1. Stochastic models
3.3.2. Deterministic models
3.3.3. Choosing between deterministic and stochastic models
4. The appropriate level of complexity for representing the system
4.1. Simplifications regarding the representation of the system
4.2. Advantages and disadvantages of using simple or complicated models
4.3. The amount of effort that one is willing to put into parameter estimation
5. General approaches to using deterministic models based on differential equations
5.1. Kinetics of the hydrolysis of linear homopolysaccharides
5.2. Is it reasonable to treat reactions as pseudo-first order in substrate concentration?
5.3. How to describe processivity in deterministic models?
6. General approaches to using stochastic models
6.1. How many molecules should a stochastic simulation involve?
6.2. How to translate between numbers of molecules and concentrations?
6.3. How to model systems in which there is only one type of enzyme?
6.4. How to model systems in which there is more than one type of enzyme?
7. ``Fingerprinting models´´ as tools for estimating specificity constants
7.1. General description of the fingerprinting method
7.2. Case study to demonstrate the principles of the fingerprinting method
7.3. Considerations about the fingerprinting method
8. Conclusion
References
Chapter 12: Polysaccharide deconstruction products: Production of bio-based building blocks
1. Introduction
2. Succinic acid as a promising bio-based building block
3. Bio-based lactic acid: An important building block in biorefinery concept
4. Microbial propionic acid production
5. Conclusions
Acknowledgments
1IntroductionThe most abundant and renewable material in the world is the lignocellulosic biomass, and its fractionation is cr
References
Chapter 13: Polysaccharide degradation for oligosaccharide production with nutraceutical potential for the food industry
1. Introduction
2. Functional oligosaccharides
2.1. Properties and food industrial application
2.2. Polysaccharides sources and production process
3. Sucrose-related oligosaccharides
3.1. Fructooligosaccharides
3.1.1. Sucrose as substrate
3.1.2. Inulin as substrate
4. Lactose-related oligosaccharides
4.1. Galactooligosaccharides, lactulose, and lactosucrose
5. Starch-related oligosaccharides
5.1. Malto-oligosaccharides
5.2. Trehalose
5.3. Isomalto-oligosaccharides
5.4. Cyclodextrins
6. Nonstarch oligosaccharides
6.1. Xylo-oligosaccharides
6.2. Cello-oligosaccharides
6.3. Pectin-oligosaccharides
6.4. Soy-oligosaccharides
6.5. Chitosan-oligosaccharides
7. Algal-oligosaccharides
7.1. Agaro and neoagaro-oligosaccharides
7.2. Alginate-oligosaccharides
7.3. Carrageenan-oligosaccharides
7.4. Fucoidan-oligosaccharides
7.5. Laminarin-oligosaccharides
7.6. Porphyran-oligosaccharides
8. Concluding remarks
Acknowledgment
References
Chapter 14: Carbohydrate-active enzymes in the production of lactose-derived tagatose
1. d-Tagatose and production strategies of a rare sugar
1.1. Whey: An essential substrate for the production of tagatose
2. β-Galactosidase and its applications
3. l-Arabinose isomerase
4. Integrated production of tagatose using immobilized enzymes
References
Chapter 15: Immobilized biocatalysts for hydrolysis of polysaccharides
1. Introduction
2. Enzyme immobilization
2.1. Fundamentals of enzyme immobilization
2.2. Techniques and materials
2.2.1. Enzyme cross-linking
2.2.2. Enzyme encapsulation or entrapment
2.2.3. Biding enzymes to supports
3. Materials and techniques applied to hydrolysis of polysaccharides
4. Industrial applications
4.1. Food industry
4.2. Textile industry
4.3. Human nutrition
4.3.1. Lactose intolerance
4.3.2. Prebiotics
4.4. Animal nutrition
4.5. Pharmaceutical industry
4.5.1. Monoclonal antibodies
4.5.2. Oral drug delivery
4.6. Water treatment
4.6.1. Removal of pollutants
4.6.2. Removal of pathogens
4.7. Energy demand
5. Final considerations
Acknowledgments
References
Chapter 16: Carbohydrate-based economy: Perspectives and challenges
1. Introduction
2. Market opportunities for carbohydrate-based products
2.1. Circular bioeconomy: Waste valuation for biofuel production
2.2. Carbohydrate-based chemicals, materials, and devices
2.2.1. Chemical modification of carbohydrate-based raw materials
2.2.2. Carbohydrate-containing nanoproducts
2.2.3. Composites and other carbohydrate-combined products
2.3. Industrial enzymatic biocatalysis
2.3.1. Food and beverage industries
2.3.2. Textile industries
2.4. Bioactive fungi compounds from carbohydrate sources aimed at pathogen control
2.4.1. Antiprotozoal activity
2.4.2. Antibacterial
2.4.3. Antiviral
3. What about the operational and environmental point of view?
References
Index
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