Transcript Industrial Production & Bioremediation

Industrial Production &
Bioremediation
Microbes for industrial production
Preservation of cultures
Methods of industrial production
Major products of industrial microbiology
Bioremediation
Biosensors & microarrays
Microbes for industrial production
Finding microorganisms in nature
 Only a small percentage of microbial species
have been cultured
 Bioprospecting: Hunting for new
microorganisms with potential for commercial
exploitation
 Great deal of interest in microbes from extreme
environments
 Challenge is to develop cost-effective
techniques for their culture
Microbes for industrial production
Genetic manipulation
 Altering the characteristics of existing known
species to produce new and desirable
characteristics
 Mutations can be induced with mutagenic
agents or UV irradiation
 Example: Development of high-yield cultures of
Penicillium for penicillin production
 Protoplast fusion can be used to fuse cells of
eukaryotic microbes and microbes that are not
phylogenetically related; used especially for
genetic manipulation in yeasts & molds
Microbes for industrial production
Genetic manipulation
 Altering the characteristics of existing known
species to produce new and desirable
characteristics
 Mutations can be induced with mutagenic
agents or UV irradiation
 Example: Development of high-yield cultures of
Penicillium for penicillin production
 Protoplast fusion can be used to fuse cells of
eukaryotic microbes and microbes that are not
phylogenetically related; used especially for
genetic manipulation in yeasts & molds
Microbes for industrial production
Genetic manipulation
 Altering the characteristics of existing known
species to produce new and desirable
characteristics
 Mutations can be induced with mutagenic
agents or UV irradiation
 Example: Development of high-yield cultures of
Penicillium for penicillin production
 Protoplast fusion can be used to fuse cells of
eukaryotic microbes and microbes that are not
phylogenetically related; used especially for
genetic manipulation in yeasts & molds
Microbes for industrial production
Genetic manipulation
 Altering the characteristics of existing known
species to produce new and desirable
characteristics
 Mutations can be induced with mutagenic
agents or UV irradiation
 Example: Development of high-yield cultures of
Penicillium for penicillin production
 Protoplast fusion can be used to fuse cells of
eukaryotic microbes and microbes that are not
phylogenetically related; used especially for
genetic manipulation in yeasts & molds
Microbes for industrial production
Genetic manipulation
 Altering the characteristics of existing known
species to produce new and desirable
characteristics
 Mutations can be induced with mutagenic
agents or UV irradiation
 Example: Development of high-yield cultures of
Penicillium for penicillin production
 Protoplast fusion can be used to fuse cells of
eukaryotic microbes and microbes that are not
phylogenetically related; used especially for
genetic manipulation in yeasts & molds
Microbes for industrial production
Genetic manipulation
 Altering the characteristics of existing known
species to produce new and desirable
characteristics
 Mutations can be induced with mutagenic
agents or UV irradiation
 Example: Development of high-yield cultures of
Penicillium for penicillin production
 Protoplast fusion can be used to fuse cells of
eukaryotic microbes and microbes that are not
phylogenetically related; used especially for
genetic manipulation in yeasts & molds
Microbes for industrial production
Genetic manipulation
 Site-directed mutagenesis is the insertion of
short segments of DNA (using recombinant
DNA technology) into a gene to lead to desired
changes in its protein product
 Recombinant DNA can be transferred between
different organisms, creating combinations of
genes with exhibit desired characteristics
 Shuttle vectors: Vectors (such as bacterial plasmids)
that can replicate in more than one species
 Expression vectors: Vectors that have transcriptional
promoters capable of mediating gene expression in
the target species.
Microbes for industrial production
Genetic manipulation
 Gene expression can be modified by altering
transcriptional regulation, fusing proteins, and
removing feedback regulation controls
 This is used for pathway architecture, or metabolic
pathway engineering, to increase or regulate
production.
 Natural genetic engineering
 Growing cultures under marginal (“stressful”)
growth conditions and selecting for new strains
(spontaneous mutations) that have increased growth
in those conditions
Preservation of cultures
Periodic transfer + refrigeration
Mineral oil slant + refrigeration
Washed culture + refrigeration
Freezing
Freezing with 50% glycerol
Drying
Lyophilization (freeze drying)
Ultracold freezing
Methods of industrial production
Medium development
 Lower-cost ingredients, such as crude plant or
animal by-products, are used for costeffectiveness
 Manipulating the levels of a limiting nutrient
may be critical to trigger or optimize the
production of a desired product
Methods of industrial production
Scaleup
 Successive optimization of growth & product
yield from a small scale (such as a shaking
flask or small fermenter) to a large scale (such
as industrial scale fermenters)
 Mixing, aeration, pH control, foaming, &
formation of filamentous growth or biofilms are
significant issues in scaleup
Methods of industrial production
Methods for mass culture
 Batch fermentation
 Continuous culture (chemostat)
 Lift-tube fermentation
 Solid-state fermentation
 Fixed-bed reactors
 Fluidized-bed reactor
 Dialysis culture unit
Methods of industrial production
Primary & secondary metabolites
 Primary metabolites are produced during the
growth phase of the microbe. Examples: amino
acids, nucleotides, fermentation end products,
and many types of enzymes
 Secondary metabolites accumulate during
periods of nutrient limitation and waste
buildup. Examples: many antibiotics and
mycotoxins
Major products
Antibiotics
 Examples: penicillin & streptomycin
 The yield of both of these antibiotics are
optimized by nutrient limitation (carbon &
nitrogen)
Recombinant DNA products
 Proteins produced from genes introduced into
microbes via recombinant DNA techniques,
such as enzymes, peptide hormones,
recombinant vaccines
Major products
Amino acids
 Glutamic acid (monosodium glutamate) is
produced by regulatory mutants of
Corynebacterium glutamicum that have a
modified Krebs cycle that can be manipulated
to shift -ketoglutarate to glutamate
production
 Lysine is produced by a Corynebacterium
glutamicum strain in which homoserine lactone
synthesis is blocked
Major products
Other organic acids
 Acetic acid, citric acid, fumaric acid, gluconic
acid, itaconic acid, kojic acid, lactic acid
“Speciality” compounds
 A variety of drugs (cholesterol drugs,
immunosuppressants, antitumor drugs),
ionophores, enzyme inhibitors, pesticides
Biopolymers
 Microbial-produced polymers, mostly
polysaccharides, useful as thickening or gelling
agents in foods, pharmaceuticals, paints, etc.
Major products
Biosurfactants
 Microbial-produced detergents, such as
glycolipids; used in bioremediation applications
such as oil spill cleanups
Bioconversions
 Using a microbe as a biocatalyst to convert a
substrate into a desired product; for example, in
the modification of steroid hormones
Bioremediation
Biodegradion in natural communities
 Includes:
 minor changes in organic molecules, leaving the
main structure still intact
 fragmentation of an organic molecule into smaller
organic molecules, still resembling the original
structure
 complete mineralization of an organic molecule to
CO2
 Recalcitrant compounds are organic compounds
that are resistant to biodegradation
Bioremediation
Biodegradion in natural communities
 Halogenated compounds, especially
halogenated aromatic compounds (such as
polychlorinated biphenyls) are often recalcitrant
 The presence of halogens in a meta position
makes the compound more recalcitrant
 Often one stereoisomer of an organic
compound will be biodegradable, while another
isomer will be recalcitrant
 Specific organisms in an environment may be
able to degrade recalcitrant compounds, at
varying rates depending on the conditions
Bioremediation
Biodegradation in natural communities
 Sometimes partial degradation of a compound
may yield compounds that are worse; for
example, trichloroethylene can be degraded to
form highly carcinogenic vinyl chloride
 Another example of detrimental biodegradation
is microbial corrosion of metal pipes
Bioremediation
Stimulating biodegradation
 Biodegradation by naturally-occurring
organisms may be stimulated by
 Adding essential nutrients to the contaminated area
 Providing aeration or limiting aeration, depending
on whether the contamination is better degraded
under aerobic or anaerobic conditions
 Using plants and the microbial communities of their
rhizospheres (phytoremediation)
 Using microbes for metal bioleaching from
minerals
Bioremediation
Bioaugmentation
 Adding microbes not normally found in an
environment to try to alter or accelerate the
biodegradation process
 When the microbes are added without
consideration of their “normal” habitat (e.g.,
just adding a pure culture), there may be shortterm improvement but the added microbe
usually fails to establish a stable population
 Better results are may be seen when the added
organism’s microenvironment (nutrients,
oxygen, aeration, etc.) are included in the
bioaugmentation strategy
Biosensors & microarrays
Biosensors
 Devices in which a biospecific molecule (e.g., a
monoclonal antibody or a hormone receptor
protein) is attached to a “transducer” (often a
piezoelectrically-active quartz chip)
 When the biosensor binds to its target, it slighty
“twists” the transducer, creating a small
electrical current that can be amplified,
detected, and measured
Biosensors & microarrays
Microarrays
 A series of microscopic DNA spots on a glass,
plastic, or silicon backing; used to monitor
levels of gene expression for thousands of
genes simultaneously, or to determine
differences in genotype