Microbial Production of Hydrogen

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Microbial Production of Hydrogen

• Jeff Tuckis
• ABE 6663

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Outline

• Importance of hydrogen production
• Types of hydrogen production
• Fermentative
• Photosynthetic
• The hydrogenase enzyme
• Hydrogen production by Chlamydomonas reinhardtii
• Research proposal

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Importance of Hydrogen as an Alternative Fuel

• Increased levels of CO2 from fossil fuels cause
  an increase in the Greenhouse Effect
• One of the detrimental effects of the Greenhouse
  Effect is Global Warming
• Combustion of Hydrogen produces water, which is
  not detrimental to the environment

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Effects of Global Warming

• Changes in climate
• Sea level rise due to melting of polar icecaps
• US seas rising 2.5-3.0mm/yr
• By year 2050 it is most likely sea levels will
  rise by 15cm, but 1 chance they will rise one
  meter (Titus et al 1991)
• One meter rise in sea level will cause 270-475
  billion dollars in damage (Titus and Narayanam
  1995)

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Microbial Hydrogen Production

• Types of microbial hydrogen production
• Fermentative
• Photosynthetic (aerobic/anaerobic)
• Most interest in hydrogen production research in
  US during the Energy Crisis of the 1970s
• Interest in hydrogen production again in 1990s
  due to the awareness of Global Warming, etc.

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Fermentative Production of Hydrogen

• Clostridia species - Clostridia beijerincki
• Used in fuel cell that produced 15mA over 20 days
  using waste from alcohol distillery (Taguchi et
  al 1992)
• Methanogens -Methanotrix soehngenii
• Archeabacteria -Pyrococcus furiosus
  (hyperthermophile)
• Eschericia coli - Formate Hydrogen Lyase pathway,
  which is inefficient

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Photosynthetic Production of Hydrogen

• Purple Sulfur bacteria (Thiocapsa and
  Chromantium)
• Non-Sulfur bacteria (Rhodospirillum and
  Rhodopseudomonas)
• Green Algae (Chlamydomonas reinhardtii)
• Advantages
• Photosynthetic organsims produce more hydrogen
  than fermentative organisms (Nandi and Sengupta
  1998)
• Photosynthetic organisms only require light and
  water

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Photosynthetic Hydrogen Production Mechanisms

• The purple sulfur bacteria and the non-sulfur
  bacteria produce hydrogen through a reversible
  hydrogenase and as a by-product of
  denitrification.

• Chlamydomonas reinhardtii a green algae, produces
  hydrogen through hydrogenase.
• Hydrogenase is induced under anaerobic conditions

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Hydrogenase

• Many bacteria use it for H2 dissimulation to use
  the electrons for electron transport
• Two types of hydrogenases
• Nickel-Iron centered - dissimulate hydrogen
• Iron-only centered - evolve H2

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Mechanism of Hydrogenase

• Hydrogen Dissimulation
• H2 Hydrogenase 2H 2e-
• Hydrogen Evolution
• Fd-reduced 2H Hydrogenase H2

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Comparison of Hydrogenase Yields
Chlamydomonas has the highest hydrogen yield
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Advantages of Using C. reinhardtii

• Cheap and easy to grow
• Requires fluorescent light and 5 CO2
• Grows at room temp. in water
• Mutants have been isolated that are more
  energetically efficient (lack Photosystem I)
• Research into its life cycle and flagella have
  yielded useful molecular techniques for studying
  the organism

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Chlamydomonas reinhardtii

• Green-alga (eukaryotic)
• Biflagellated, unicellular, photosynthetic
• Reproduces asexually or sexually under adverse
  conditions

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Chlamydomonas reinhardtii
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C. reinhardtii Hydrogenase

• Enzyme located in the chloroplast
• Receives electrons from reduced ferrodoxin
• Hydrogenase stimulated under anaerobic conditions
  (Happe et el 1994)
• Hydrogenase inhibited by O2, which is produced by
  C. reinhardtii during photosynthesis

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Research Needs

• Investigate hydrogenase oxygen inhibition
• Investigate production of hydrogen by the
  Photosystem I mutant

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Research Proposal

• Problem Hydrogenase in C. reinhardtii inhibited
  by oxygen, which is made by the organism.
• Solution Determine Oxygen binding site on
  hydrogenase and create a clone with lower O2
  binding and therefore higher H2 production.

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Research Approach

• Isolate gene for hydrogenase (BAC library has
  recently become available)
• Sequence the hydrogenase gene and determine the
  oxygen binding site.
• Create hydrogenase clone that does not bind
  oxygen through site-directed mutagenesis.
• Isolate hydrogenase deficient C. reinhardtii
  mutant through UV-mutagenesis.
• Introduce mutant hydrogenase into hydrogenase
  deficient clone.