Current Developments in Biotechnology and Bioengineering: Designer Microbial Cell Factories: Metabolic Engineering and Applications

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Current Developments in Biotechnology and Bioengineering
Copyright Page
Contents
List of contributors
Preface
I. Metabolic Engineering of Cells: General and Basics
	1 Metabolic engineering: tools for pathway rewiring and value creation
		1.1 Introduction
		1.2 Tools for metabolic engineering
			1.2.1 Microbial strain selection and improvement
			1.2.2 Synthetic biology tools
			1.2.3 Protein engineering tools
			1.2.4 Omics tools
				1.2.4.1 Genomics and transcriptomics tools
				1.2.4.2 Proteomics and metabolomics tools
				1.2.4.3 Fluxomics tools
				1.2.4.4 Meta-omics tools
			1.2.5 Genome engineering tools
		1.3 Value generation by metabolic engineering
		1.4 Conclusions and perspectives
		References
	2 Membrane transport as a target for metabolic engineering
		2.1 Introduction
		2.2 Membrane transport proteins
		2.3 Substrate uptake
		2.4 Transport from and into organelles
		2.5 Product export
		2.6 Cellular robustness
		2.7 Substrate channeling and membrane transport
		2.8 Undesired transport processes
		2.9 Conclusions and perspectives
		References
	3 Analysis and modeling tools of metabolic flux
		3.1 Introduction
		3.2 13C-metabolic flux analysis
			3.2.1 13C-metabolic flux analysis in steady-state systems
			3.2.2 13C-metabolic flux analysis in nonstandard systems
				3.2.2.1 13C-metabolic flux analysis of autotrophic metabolism
				3.2.2.2 13C-metabolic flux analysis in the nongrowth stage
				3.2.2.3 13C dynamic metabolic flux analysis
		3.3 Constraint-based stoichiometric metabolic flux analysis
			3.3.1 Genome-scale metabolic network model tools
				3.3.1.1 Method of genome-scale metabolic network model construction
					3.3.1.1.1 Manual model reconstruction method
					3.3.1.1.2 Automatic model reconstruction method
				3.3.1.2 Database and software required for model construction
				3.3.1.3 Model analyzing algorithm
					3.3.1.3.1 Flux balance analyzing algorithm
					3.3.1.3.2 Genetic disturbance algorithm
			3.3.2 Research progress of genome-scale metabolic network model
				3.3.2.1 Model reconstruction
				3.3.2.2 Model application
					3.3.2.2.1 Predict and analyze the growing phenotype of microorganisms
					3.3.2.2.2 Analysis of network properties
					3.3.2.2.3 Guide metabolic engineering
		3.4 Conclusions and perspectives
		References
	4 Equipped C1 chemical assimilation pathway in engineering Escherichia coli
		4.1 Introduction
		4.2 Approaches for the assessment of CO2 assimilation capability
		4.3 Physiological effect of RuBisCo system
		4.4 Strategies to enhance the RuBisCo system
		4.5 Transforming the heterotrophs to autotrophs
		4.6 Prospective of RuBisCo-based chemical production
		4.7 Conclusions and perspectives
		References
	5 Microbial tolerance in metabolic engineering
		5.1 Introduction
		5.2 Microbial stresses and responses
			5.2.1 Physiological stress factors
				5.2.1.1 Chemical stresses
				5.2.1.2 Physical stresses
			5.2.2 Microbial responses to stress factors
				5.2.2.1 Regulatory responses to stress factors
				5.2.2.2 Removal of the toxic chemicals or metabolic intermediates by enzymatic conversion
				5.2.2.3 Alteration of membrane structure and/or transport
				5.2.2.4 Protein homeostasis
				5.2.2.5 Cross tolerance
		5.3 Strategies to improve microbial tolerance
			5.3.1 Rational approaches
				5.3.1.1 Expression of efflux pump
				5.3.1.2 Overexpression of chaperones
				5.3.1.3 Engineering of cell envelope
				5.3.1.4 Engineering of transcriptional regulatory systems
			5.3.2 Adaptive laboratory evolution
				5.3.2.1 Adaptive laboratory evolution for tolerance against organic acids and alcohols
				5.3.2.2 Adaptive laboratory evolution for tolerance against salts and dissolved oxygen
				5.3.2.3 Adaptive laboratory evolution and system-level analyses
		5.4 Challenges in developing and using tolerant strains
		5.5 Conclusions and perspectives
		References
	6 Application of proteomics and metabolomics in microbiology research
		6.1 Introduction
		6.2 Proteomics in microbiology
			6.2.1 Proteomic methodology
				6.2.1.1 Bottom-up proteomics
					Discovery and targeted proteomics
					Label and label-free quantification
				6.2.1.2 Top-down proteomics
			6.2.2 Proteomic application in microbiology
				6.2.2.1 Identification of pathogenic bacteria
				6.2.2.2 Host-pathogen interactions
				6.2.2.3 Characterization of biological membrane
				6.2.2.4 Antibiotics resistance
				6.2.2.5 Advanced growth rate of bacteria
				6.2.2.6 Energy conversion
		6.3 Metabolomics in microbiology
			6.3.1 Metabolomic methodology
				6.3.1.1 Nuclear magnetic resonance spectroscopy
				6.3.1.2 Mass spectrometer
					Gas chromatography-MS
					Liquid-chromatography-MS
				6.3.1.3 Metabolites identification
			6.3.2 Metabolomic applications in microbiology
				6.3.2.1 Improvement of the production of various bioproduction targets
				6.3.2.2 Exploration of the effect of unknown gene
				6.3.2.3 Development of synthetic methylotrophic strains
		6.4 Conclusions and perspectives
		References
	7 Approaches and tools of protein tailoring for metabolic engineering
		7.1 Introduction
		7.2 Approaches for the engineering of protein
			7.2.1 Directed evolution
			7.2.2 Rational design
			7.2.3 Semirational design or combined methods
		7.3 Applications of protein engineering
			7.3.1 Enzyme engineering for improving the catalytic activity
			7.3.2 Engineering the enzymes for cofactor utilization
			7.3.3 Regulatory protein engineering for the enhancement of the metabolic pathway
			7.3.4 Altering substrate and product specificity
			7.3.5 Engineering the regulatory elements of the enzymes
			7.3.6 Assimilation of unnatural amino acid into a protein
			7.3.7 Enzymes scaffold engineering to control metabolite flux
			7.3.8 De novo engineering of enzymes
			7.3.9 Formation of enzyme complex through colocalization for the advancement in the metabolic pathway
		7.4 Conclusions and perspectives
		References
	8 Microbial metabolism of aromatic pollutants: High-throughput OMICS and metabolic engineering for efficient bioremediation
		8.1 Introduction
		8.2 Aromatic compounds: impact and toxicity
		8.3 Microbial metabolism of aromatic compounds/pollutants
			8.3.1 Microbes involved
			8.3.2 Metabolic pathways
			8.3.3 Enzymes involved in the metabolism
		8.4 High-throughput OMICS: insights into aromatics metabolism
			8.4.1 Metagenomics
			8.4.2 Metatranscriptomics
			8.4.3 Metaproteomics
			8.4.4 Meta-metabolomics
		8.5 Metabolic engineering for efficient aromatics biodegradation
			8.5.1 Designing the metabolic pathway
				8.5.1.1 Pathway prediction tools
				8.5.1.2 Toxicity prediction databases
				8.5.1.3 Molecular biology module databases
				8.5.1.4 Metabolic modeling
				8.5.1.5 Choosing the ideal chassis (host)
			8.5.2 Building the desired strain
				8.5.2.1 Plasmid-based expression
				8.5.2.2 Genome engineering strategies
					Homologous recombination
					Recombineering
					Transposon-based tools
			8.5.3 Testing and analyzing the engineered strain
		8.6 Conclusions and perspectives
		References
	9 Microbial consortium engineering for the improvement of biochemicals production
		9.1 Introduction
		9.2 Classification of microbial consortia
			9.2.1 Modes of construction
				9.2.1.1 Natural microbial consortium
				9.2.1.2 Artificial microbial consortium
				9.2.1.3 Synthetic microbial consortium
			9.2.2 Modes of interaction
				9.2.2.1 Synergistic
				9.2.2.2 Mutualistic
				9.2.2.3 Commensalism
				9.2.2.4 Competition
				9.2.2.5 Parasitism
			9.2.3 Functional modes
				9.2.3.1 Environment maintenance consortium
				9.2.3.2 Nutrient exchange consortium
				9.2.3.3 Substrate facilitator consortium
				9.2.3.4 Signal exchange consortium
		9.3 Construction of a microbial consortium
			9.3.1 Source of carbon
			9.3.2 Inoculum ratio
			9.3.3 Spatial organization
			9.3.4 Physiological parameters
			9.3.5 Availability of nutrients
		9.4 Applications of microbial consortium engineering
			9.4.1 Production of bioactive molecules
			9.4.2 Production of biopolymers
				9.4.2.1 Polyhydroxyalkonates
				9.4.2.2 Exopolysaccharides
			9.4.3 Production of fermented food products
			9.4.4 Bioenergy
				9.4.4.1 Biodiesel
				9.4.4.2 Bioalcohol
				9.4.4.3 Biohydrogen
			9.4.5 Production of biochemicals
				9.4.5.1 Production of acetic acid
				9.4.5.2 Production of butyric acid
				9.4.5.3 Production of carotenoids
				9.4.5.4 Production of single-cell protein
			9.4.6 Bioremediation
				9.4.6.1 Degradation of heavy metals
				9.4.6.2 Degradation and decolorization of textile effluent
				9.4.6.3 Degradation of petroleum waste
				9.4.6.4 Water eutrophication
		9.5 Recent synthetic microbial consortia and their applications
			9.5.1 Coculturing
			9.5.2 Metabolic engineering
			9.5.3 Enzyme engineering
			9.5.4 Fermentation systems
		9.6 Challenges in microbial consortium engineering
		9.7 Conclusions and perspectives
		References
		Further reading
II. Metabolic Engineering of Cells: Applications
	10 Metabolic engineering strategies for effective utilization of cellulosic sugars to produce value-added products
		10.1 Introduction
		10.2 Sustainable carbon sources for biorefineries
			10.2.1 Carbohydrates from lignocellulosic biomass
				10.2.1.1 Cellulose and glucose
				10.2.1.2 Hemicellulose and xylose
				10.2.1.3 Cellobiose and other cellodextrins
			10.2.2 Levulinic acid from cellulose
		10.3 Microbial cell factories for carbon source coutilization and production of value-added chemicals
			10.3.1 Escherichia coli
			10.3.2 Corynebacterium glutamicum
			10.3.3 Pseudomonas putida
			10.3.4 Saccharomyces cerevisiae
		10.4 Conclusions and perspectives
		References
	11 Production of fine chemicals from renewable feedstocks through the engineering of artificial enzyme cascades
		11.1 Introduction
		11.2 Advantages of enzyme cascades
		11.3 Artificial enzyme cascades versus natural enzyme cascades
		11.4 Importance of fine chemicals production from renewable feedstocks through artificial enzyme cascades
		11.5 General principle of engineering of enzyme cascades
			11.5.1 Basic designs of cascade
				11.5.1.1 Linear cascades
				11.5.1.2 Orthogonal cascades
				11.5.1.3 Cyclic cascades
				11.5.1.4 Coupled cascades
				11.5.1.5 Divergent cascades
			11.5.2 Operation of enzyme cascade reactions
				11.5.2.1 In vitro cascade reaction
				11.5.2.2 In vivo cascade reaction
			11.5.3 The development of enzyme cascades for the conversion of renewable feedstocks to high-value fine chemicals
				11.5.3.1 Pathway design
				11.5.3.2 Enzyme selection
				11.5.3.3 Engineering strains for expressing all enzymes required in cascade
		11.6 Examples of production of fine chemicals from bio-based l-phenylalanine using artificial enzyme cascades
			11.6.1 Artificial enzyme cascades for the production of alcohols
			11.6.2 Artificial enzyme cascades for the production of carboxylic acids
			11.6.3 Artificial enzyme cascades for production of amines
		11.7 Examples of production of fine chemicals from renewable feedstocks glucose and glycerol using artificial enzyme cascades
			11.7.1 Single strain approach
			11.7.2 Coupled strains approach
		11.8 Conclusions and perspectives
		References
	12 Metabolic engineering of microorganisms for the production of carotenoids, flavonoids, and functional polysaccharides
		12.1 Introduction
		12.2 Metabolic engineering of plant natural products
			12.2.1 Flavanones
			12.2.2 Flavones
			12.2.3 Flavonols
			12.2.4 Flavanols
			12.2.5 Anthocyanins
			12.2.6 Isoflavones
			12.2.7 Resveratrol
			12.2.8 Limonene
			12.2.9 Lycopene
			12.2.10 Carotene
			12.2.11 Astaxanthin
			12.2.12 β-Ionone
			12.2.13 Emodin
			12.2.14 Cannabinoids
		12.3 Metabolic engineering of functional polysaccharides
			12.3.1 Levan
			12.3.2 Hyaluronic acid
			12.3.3 Heparosan and chondroitin
		12.4 Conclusions and perspectives
		References
	13 Bioengineering in microbial production of biobutanol from renewable resources
		13.1 Introduction
		13.2 Applications and production of butanol
		13.3 Biological production of butanol
		13.4 Metabolic pathways of biobutanol production
			13.4.1 Acetone–butanol–ethanol pathway
			13.4.2 Keto-acid pathways
		13.5 Enhancement of biobutanol production
			13.5.1 Optimization of fermentation conditions
			13.5.2 Bioengineering for biobutanol production
				13.5.2.1 Spectrum of fermentation substrates
				13.5.2.2 Modified pathways
				13.5.2.3 Increased solvent tolerance
		13.6 Conclusions and perspectives
		References
	14 Engineered microorganisms for bioremediation
		14.1 Introduction
		14.2 Types of bioremediation
			14.2.1 Limitations associated with bioremediation
		14.3 Genetically engineered organisms in bioremediation
			14.3.1 Genetically modified bacteria in bioremediation
			14.3.2 Genetically modified fungi in mycoremediation
			14.3.3 Algae in phycoremediation
			14.3.4 Genetically modified plants in phytoremediation as an alternative
		14.4 Genetic engineering techniques
			14.4.1 Protein engineering
			14.4.2 Pathway modification
			14.4.3 Advanced genome engineering
			14.4.4 Quorum sensing: an emerging area for bioremediation
		14.5 Bioremediation using GEMs
			14.5.1 Heavy metals
			14.5.2 Pesticides and herbicides
			14.5.3 Dyestuff
			14.5.4 Oils and petroleum products
		14.6 Field applications of GEMs
			14.6.1 Bioremediation potential
			14.6.2 Survival in harsh habitats
			14.6.3 Bioprocess controlled monitoring
		14.7 Risk assessment of GEMs
		14.8 Conclusions and perspectives
		References
	15 Agricultural applications of engineered microbes
		15.1 Introduction
		15.2 Agricultural applications of genetically modified microbes
			15.2.1 Applications of genetically modified microbes in plant growth and nutrition
				15.2.1.1 Symbiotic nitrogen fixers
					Genetically modified Rhizobium
					Genetically modified Alcaligenes faecalis
					Genetically modified Xanthobacter autotrophicus
				15.2.1.2 Nonsymbiotic nitrogen fixers
					Genetically modified Azospirillum
				15.2.1.3 Genetically modified phosphate-solubilizing microbes
			15.2.2 Applications of genetically modified microbes in plant stress tolerance
				15.2.2.1 Role of genetically modified microbes to control insect pests
				15.2.2.2 Role of genetically modified microbes to control plant diseases
				15.2.2.3 Application of genetically modified microbes in plant protection from ice-nucleation
		15.3 Conclusions and perspectives
		References
	16 Rhizosphere microbiome engineering
		16.1 Introduction
		16.2 Plant-associated microbes/microbiome
			16.2.1 Holobiome
			16.2.2 Plant growth-promoting rhizobacteria and their contribution in plant growth and development
				16.2.2.1 Enhancement of plant nutrient acquisition
				16.2.2.2 Dealing with abiotic stress
				16.2.2.3 Phytohormone production
				16.2.2.4 Antagonism against phytopathogens
			16.2.3 Need for rhizosphere engineering
		16.3 Rhizosphere microbiome engineering
			16.3.1 Soil amendment
				16.3.1.1 Inorganic/organic soil amendment
				16.3.1.2 Microbial inoculants as biofertilizer/biopesticide
			16.3.2 Targeted plant-microbe engineering
		16.4 Emerging areas of research
			16.4.1 Organic amendments/root exudates: biochemical approach
				16.4.1.1 Movement toward rhizosphere
				16.4.1.2 Survival within the rhizosphere
				16.4.1.3 Adhesion and colonization on root surfaces
			16.4.2 Artificial microbial consortia
			16.4.3 Microbial-based plant breeding
			16.4.4 Host mediated microbiome engineering: molecular strategies
		16.5 Conclusions and perspectives
		References
	17 Genetically engineered microbes in micro-remediation of metals from contaminated sites
		17.1 Introduction
		17.2 Classification of bioremediation
			17.2.1 In situ bioremediation
				17.2.1.1 Bioventing
				17.2.1.2 Bioslurping
				17.2.1.3 Bioaugmentation
				17.2.1.4 Biosorption
				17.2.1.5 Bioaccumulation
			17.2.2 Intrinsic bioremediation
			17.2.3 Engineered in situ bioremediation
			17.2.4 Ex situ bioremediation
			17.2.5 Microbes assisted bioremediation (microremediation)
		17.3 Metal-contaminated sites: a problem
			17.3.1 Heavy metal remediation
			17.3.2 Heavy metals and toxicity
			17.3.3 Microbial biorecovery of heavy metals
		17.4 Genetically modified micro-organisms
			17.4.1 Wild type microbes
			17.4.2 Engineered microbes
				17.4.2.1 Mercury
				17.4.2.2 Arsenic
				17.4.2.3 Lead and cadmium
		17.5 Conclusions and perspectives
		References
	18 Biofuel production from renewable feedstocks: Progress through metabolic engineering
		18.1 Introduction
		18.2 Heterologous genetic expression in plants to improve feedstock properties
		18.3 System metabolic engineering for biofuels production
			18.3.1 In silico approaches in metabolic engineering
				18.3.1.1 “Omics-”approach in system biology
				18.3.1.2 Genome-scale metabolic models
				18.3.1.3 In silico optimization methods
			18.3.2 Metabolic engineering strategies
				18.3.2.1 Overexpression of genes
				18.3.2.2 Engineering of the regulatory regions
				18.3.2.3 Knock-down/-out of competing pathways
				18.3.2.4 Directed evolution/mutations toward improving rate-limiting step
		18.4 Microbial production of biofuels from renewable feedstock
			18.4.1 Bioethanol
			18.4.2 Biodiesel
				18.4.2.1 Biodiesel from microbes
				18.4.2.2 Algal biodiesel
			18.4.3 Drop-in biofuels
		18.5 Challenges and techno-economic analysis of emerging biofuels
		18.6 Conclusions and perspectives
		References
	19 Synthetic biology and the regulatory roadmap for the commercialization of designer microbes
		19.1 Introduction
		19.2 Synthetic biology
		19.3 Framework of synthetic biology
		19.4 Tools in synthetic biology
			19.4.1 Design of gene circuit
			19.4.2 Synthetic transcription factors
			19.4.3 Genome engineering
			19.4.4 Computer-aided tools in synthetic biology
		19.5 Applications of synthetic biology
			19.5.1 Production of advance biofuels
			19.5.2 Applications in drug development
			19.5.3 Bioremediation of pollutants with integrated synthetic biology
		19.6 Legal aspect of designer microbes
			19.6.1 Mentioned below are the federal laws, regulations and policies
				19.6.1.1 Europe
				19.6.1.2 United States of America (USA)
				19.6.1.3 India
				19.6.1.4 China
				19.6.1.5 Japan
			19.6.2 Other regulations
				19.6.2.1 The cartagena protocol on biosafety to the convention on biological diversity
				19.6.2.2 The codex alimentarius commission (Codex)
		19.7 Regulatory challenges for the commercialization of designer microbes
			19.7.1 Social and economic issues
			19.7.2 Biosafety issues
			19.7.3 Other issues
			19.7.4 Risk assessment and management
		19.8 Conclusions and perspectives
		References
Index
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