Metabolic Engineering: Overview, Relevance, and Future Applications

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Metabolic EngineeringOverview, Relevance, and
Future Applications

• James R. Foster II
• ABE4660
• Spring 2003

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What is metabolic engineering?

• The directed modification of cellular metabolism
  and properties through induction, deletion and/or
  modification of metabolic pathways by using
  recombinant DNA and other microbiological
  techniques Lee 1.
• Basically, the goal is to predict how changes in
  a cells environment/nutrients will affect
  existing metabolic pathways. If we can predict
  how changes will affect pathways, we can
  specifically modify an organism to produce a
  useful, profitable product

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How is metabolic engineering industrially
relevant? (2 slides)

• 5 Main Areas
• Enhanced production of metabolites or
  host-synthesized materials.
• Example Ethanol, phenylalanine
• Production of NEW metabolites
• Example E. Coli production of polyhydroxyalkanoat
  es (biodegradable polymer)
• Broadening substrate utilization range
• Example Ethanol production from lactose and
  xylose by S. cerevisiae

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How is metabolic engineering industrially
relevant? (2 of 2)

• Improving or designing new metabolic pathways for
  the degradation of chemicals.
• Example Benzene, toluene degradation by
  Pseudomonas putida
• Modification of cell properties that facilitate
  bioprocessing
• Example Better growth of E. Coli under aerobic
  conditions

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What steps are involved?
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Engineering an organism to produce a specific
product.

• Qualities in a suitable organism
• KEY POINT Metabolic pathways must be well
  understood!
• Capable of growth to high organism density (cells
  per unit area)
• Easy maintenance of cell culture
• Sequenced genome
• Common organisms
• E. coli
• S. cerevisiae
• Other microorganisms with specialized pathways

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Engineering an organism to produce a specific
product.

• Once an organism is chosen, how do we modify it?
• 1. Channel nutrients down existing metabolic
  pathways that produce the desired product.
• 2. If option 1 is not possible, use recombinant
  DNA to actually change genome and create NEW
  metabolic pathways

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Channeling nutrients down specific pathways

• Goal
• Restrict or add excess of specified nutrients for
  the purpose of inhibiting unwanted reactions and
  driving forward desired reactions
• Problem
• Nature has designed organisms with strict
  metabolic regulating mechanisms.
• Solution
• Knowledge of major and minor metabolic pathways
  aid in determining what variables/nutrients can
  be changed and how to go about changing them

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Recombinant DNA Technology

• Some organisms require special feedstocks or
  environments
• Transfer genes responsible for producing desired
  product from host organisms to convenient
  organism such as E. coli

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Recombinant DNA

• Advantages of using recombinant DNA
• Allows for choosing of host organism
• Inexpensive feedstocks
• High organism density (good product yields)
• Removes many pathways of inhibition
• Since metabolic pathways are under strict
  regulation, selectively transferring only the
  genes of interest will leave behind genes
  responsible for pathway inhibition. This results
  in greater expression of the gene.

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Areas of Research

• Biodegradable Polymers
• PHA (Polyhydroxyalkanoates)
• Multiple organisms involved
• Convenient pathways use Acetyl-CoA as carbon
  source
• However, knowledge of these organisms is
  relatively limited
• AND, many species require special feed-stocks
• Consider using E. Coli for producing PHA
• Requires use of recombinant DNA

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PHA Production
Source http//mbel.kaist.ac.kr/research/metaeng_e
n.html
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What Happens in the lab?
Poly(3-hydroxybutyrate) Production
Source http//mbel.kaist.ac.kr/publication/int40.
pdf
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Amino Acid Production

• Second largest fermentation industry
• (1st is ethanol production)
• In 1957, it was found that preventing biotin
  intake in C. glutamicum resulted in excess
  secretion of L-glutamic acid.
• Way to make monosodium glutamate
• Now all amino acids can be synthesized by
  manipulation of various pathways in
  microorganisms.

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Example of Amino Acid Production Pathway
Source http//mbel.kaist.ac.kr/research/metaeng_e
n.html
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Areas of Research

• Microbial degradation of pollutants
• Degradation of aromatic hydrocarbons
• Toluene
• Benzene
• Halogenated Compounds
• Polychlorinated biphenyls

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Source http//www.sutcliffe-croftshaw.co.uk/appl-
odour.htm
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Areas of Research

• Mammalian Cell Cultures
• Obtaining a stable culture of mammalian cells can
  be useful in the synthesis of proteins.
• Future applications
• Tissue culture
• Cancer therapy
• BUT, major problems
• Cells dependent on other cells for survival
• Apoptosis
• Solution
• Develop multi-gene systems capable of integrating
  metabolic pathways and controlling apoptosis and
  cell division

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Future of Metabolic Engineering

• Continued efforts to pioneer new methods for
  developing a green economy
• Source of energy (hydrogen gas, electrodes)
• Hazardous waste degradation
• Environmentally friendly industrial processes
• Medicine
• Protein synthesis
• Tissue/organ culturesethics?
• Cure for Cancer?......not yet
• Final Thought
• Use natures power and efficiency to benefit
  mankind and preserve the environment

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Questions?

• References
• Lee, Sang Yup. Metabolic Engineering. Marcel
  Dekker, New York, 1999.