Microbial Metabolism— Procuring Energy!

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Microbial MetabolismProcuring Energy!
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Metabolic diversity
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Key PointsDissimulative metabolismReduction of
chemicals for energymuch material must be used
to achieve sufficient energy for
growthAssimilative metabolismReduction of
chemicals for biomassthe cell only uses as much
starting material as required
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Anaerobic Respirationmolecular oxygen does NOT
serve as an electron acceptor but energy (ATP) is
produced via chemiosmosis
NO3- ? ? ? N2 or SO4-- ? H2S or CO2 ?? ? CH4
1. Nitrate ? ? ? Nitrogen gas 2. Sulfate ?
Hydrogen Sulfide gas 3. Carbon dioxide ? ? ?
methane
1,2. Standard electron transport, 3. membrane
bound enzymes Both generate proton gradient
required for PMF
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In anaerobic metabolism Nitrate or Sulfate serve
as terminal electron acceptors at the end of the
electron transport chain
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II
III
Thus under anaerobic conditions some organisms
can still obtain high levels of energy
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Mixotrophs, Energy from oxidation of organic
chemicals inorganic chemicals are reduced.
NO3- ? ? ? N2 SO4-- ? H2S
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Denitrification (Nitrate reduction)Some
Pseudomonas, Bacillus and Thiobacillus spp.
Nitrate reduction is mediated by enzymes 1.
nitrate reductase (NR) 2. nitric oxide
reductase (NcOR) 3. nitrous oxide reductase
(NsOR)
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Significance of denitrification
1. Agriculture soil nitrate that could be fixed
to ammonia and assimilated by plants are reduced
to atmospheric nitrogen and lost from the soil
(howevernitrogen fixing bacteria can restore
atmospheric nitrogen to the soil as part of the
overall nitrogen cycle)

• Acid rain atmospheric nitrous oxide is
  converted to nitric
• oxide via sunlight. This combined with nitric
  oxide released via
• denitrification reacts with ozone to form nitrite
  that returns to the
• earth as acid rain.

• Sewage treatment plants (water purification)
  Denitrifying
• bacteria are added to the sewage to convert
  nitrate to atmospheric
• nitrogen to remove nitrogen that would otherwise
  promote the
• growth of algae

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Significance of denitrification
1. Agriculture soil nitrate that could be fixed
to ammonia and assimilated by plants are reduced
to atmospheric nitrogen and lost from the soil
(howevernitrogen fixing bacteria can restore
atmospheric nitrogen to the soil as part of the
overall nitrogen cycle)
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Significance of denitrification

• Acid rain atmospheric nitrous oxide is
  converted to nitric
• oxide via sunlight. This combined with nitric
  oxide released via
• denitrification reacts with ozone to form nitrite
  that returns to the
• earth as acid rain.

NO2-
sunlight
NR
ozone
H2O
Acid rain
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Significance of denitrification
1. Agriculture soil nitrate that could be fixed
to ammonia and assimilated by plants are reduced
to atmospheric nitrogen and lost from the soil
(howevernitrogen fixing bacteria can restore
atmospheric nitrogen to the soil as part of the
overall nitrogen cycle)

• Acid rain atmospheric nitrous oxide is
  converted to nitric
• oxide via sunlight. This combined with nitric
  oxide released via
• denitrification reacts with ozone to form nitrite
  that returns to the
• earth as acid rain.

• Sewage treatment plants (water purification)
  Denitrifying
• bacteria are added to the sewage to convert
  nitrate to atmospheric
• nitrogen to remove nitrogen that would otherwise
  promote the
• growth of algae

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Sulfate ReductionBacteria responsible for this
are widespread in aquatic environments
Sulfate reduction is mediated by enzymes 1. ATP
sulfurylase (ATPS) 2. APS reductase (APSR) 3.
sulfite reductase (SR)
Excreted into environment
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Significance of sulfate reduction
Pollution of waters Sulfate reducing bacteria
are limited by the amount of organic starting
material available in the aquatic environment
which when metabolized by these bacteria provide
the electrons/protons that drive the sulfate to
sulfide reaction. Disposal of sewage and garbage
into waters provides the organic material
required for this process Sulfides are toxic to
living organisms as these sulfides combine with
iron centers of cytochromes and hemoglobin thus
inhibiting their function N.B. Fe can serve as
a detoxifying agent as they react with sulfides
to produce insoluble FeSblack sediments found
in aquatic environments are good indicators of
pollution!!!)
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Carbon dioxide reduction Methanogenesis(methanoge
ns anaerobic archaebacteria)
Complex set of reactions that take place in the
membranes of these bacteria protons for CO2
reduction come from fermentation, methanogenesis
(somewhat different from electron transport)
provides proton motive force that drives the
production of ATP
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Significance of methanogenesis
Sewage treatment plants
Insoluble sludge from primary treatment is
degraded by a variety of anaerobic bacteria
using catabolic pathways and fermentation.
(we drink)
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Anoxic digestion of sludge

• Methanogens use protons from
• catabolism and fermentation to reduce
• CO2 from fermentation to methane.
• The methane is collected as natural gas to fuel
  and heat the
• sewage plant or burned off.

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Significance of methanogenesis
Digestive processes
Methanogens are found in the rumen of cows,
sheep, deer etc. The caecum of horses and
rabbits, the large intestines of humans, cats,
dogs etc. and the hindgut of termites.

• Herbivores cows/horses/rabbits/termites
• bacteria degrade cellulose to cellobiose to
  glucose?glycolysis
• fermentation ? organic acids are assimilated and
  CO2 and H2
• are reduced to methane by methanogens
• Omnivores humans/cats/dogs/pigs vast catabolic
  processes?
• fermentation ?methanogenesis of CO2
• Methane waste Cows belch, humans expel gas and
  I dont know what the termites do!!!

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Stop here
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Chemolithotrophyobtaining energy from inorganic
chemicals1. inorganic chemicals are oxidized as
coenzymes in the electron transport chain are
reduced.2. Oxygen serves as the terminal
electron acceptor in electron transport3.
Reducing power is not derived from the catabolism
of organic matter to produce NADH and FADH
therefore these cofactors are usually not
re-oxidized in chemolithotropy4. The chemicals
that are oxidized have lower energy potential
than NADH, therefore more of these chemicals must
be oxidized to generate equivalent proton motive
force to produce ATP
Chemolithotrophs tend to grow more slowly than
chemo-organotrophs
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Chemolithotrophy
X
X
Fe2 ?Fe3 NH4 ?NO2- /NO3- H2S/S2O32- ??H2SO4
O2 ? H2O
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Sulfur and Iron oxidation(Thiobacillus
thiooxidans, Thiobacillus ferrooxidans and
others.)
1. Fe2 ?Fe3 ferrous iron to ferric iron 2.
H2S ? elemental sulfur?H2SO4 hydrogen sulfide to
sulfuric acid 3. S2O32- ??H2SO4 thiosulfate to
sulfuric acid
N.B. Ferric iron can often serve as an oxidizing
agent Sulfuric acid greatly decreases the pH of
the surrounding environment Sulfates can also be
assimilated as a food source for bacteria/plants
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Significance of sulfur and iron oxidation
Hydrothermal ventssymbiotic relationships
between animals and bacteria dwelling in these
niches (Thiobacillus, Thiomicrospora,
Thiotrix)clams, mussels, tube worms
1. Basalt/magma rich in minerals beneath ocean
floor produce cracks in ocean floor. 2.
Minerals mix with sea water and are expelled
from ocean floor. Black smokers
precipitated minerals mixed with
seawater/ 270-380oC
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Life in hydrothermal vents
There is no sunlight at these depths in the ocean
yet niches around the vents are robust with
life. Chemolithotrophs serve as primary producers

• Bacteria live in the GI tract of tube worms/ the
  gills of mussels
• and clams.
• Tube worms/mussels/clams provide CO2 as a carbon
  source for
• the bacteria.
• The bacteria oxidize hydrogen sulfide and
  thiosulfate to H2SO4
• for energy and reducing power and use this for
  the assimilation
• of CO2.
• 4. Wastes from bacterial metabolism feed the
  larger animals

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Significance of sulfur and iron oxidation
Pollution/Acid mine Drainage (Thiobacillus and
Metallogenium spp.)

• Strip coal mining exposes the pyrite
• (FeS2) in coal to oxygen.
• 2. Bacteria oxidize ferrous iron to ferric
• iron
• 3. Oxidation of sulfides to sulfuric acid
• greatly reduces the pH of the water
• 4. Ferric iron reacts with water to form
• iron III hydroxide (Fe(OH)3) which
• further lowers the pH of the water
• 5. Fe(OH)3 precipitates to form slimy
• orange coating that covers the stream bed
• (indicator for pollution)
• Acid soon kills the aquatic life at the
• bottom of stream bed

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Significance of sulfur and iron oxidation
Microbial Bioremediation Bio-leaching ? use of
bacteria to extract pure metal from ores with low
metal content Can be used to isolate almost
any divalent metal copper, uranium nickel,
cobalt, tin, zinc, etc. Insoluble metal sulfides
are oxidized to soluble metal sulfates i.e.
Copper required for electricity is in short supply
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Bioremediation
Similarly, sulfur/iron oxidizing bacteria can be
used to isolate important fuel sources. Uranium
usually found naturally as the low grade ore
uranium oxide (UO2) . Thiobacillus spp. oxidize
ferrous iron to ferric iron which In turn
oxidizes UO2 to soluble UO2SO4 Oil recovery
recovery of petroleum and hydrocarbons from oil
shales 1. Oil shales contain large amounts of
carbonates and pyrites 2. Thiobacillus oxidizes
the sulfur and iron in the pyrites to produce
acids 3. Acids dissolve the carbonates thus
increasing the porosity of the oil shales 4.
Oil can be more easily recovered from these
shales.
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Nitrification/Nitrogen oxidation by nitrifying
bacteria
A two step process mediated by two genera of
bacteria
NH4 ?NO2- Nitrosomonas spp. ammonium to
nitrite NO2- ? NO3- Nitrobacter spp. nitrite to
nitrate
N.B. 35 moles of ammonium and/or 100 moles of
nitrate are required to generate enough reducing
power and ATP to convert one mole of carbon
dioxide into organic carbon
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Significance of nitrifying bacteria
Agriculture nitrifying bacteria leach nitrogen
required by plants from the soil.
1. Positively charged Ammonium ions are absorbed
by negatively charged clay particles present in
the soil, thus retaining nitrogen 2. Negatively
charged nitrites and nitrates are not absorbed by
these clay particles and are leached into the
groundwater A. loss of nitrogen from soil B.
nitrites in the water supply are toxic 1.
nitrites combine with hemoglobin to block the
exchange of oxygen 2. nitrites react with
amino compounds to form carcinogenic
nitrosamines
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Oxidative and Anaerobic Photo-phosphorylation(ob
taining energy from sunlight)
Algae takes place in chloroplasts Cyanobacteria
occurs in thylakoid membranes Purple
bacteria/sulfur bacteria and Heliobacteria
occurs in lamellar membranes
Web sites for better understanding
http//www-micro.msb.le.ac.uk/video/photosynthesis
.html
http//www.biologie.uni-hamburg.de/b-online/chimes
/photo/ebacphot.htm
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Photosynthetic membranes of the bacteria
Lamellar membranes in a purple bacterium These
membranes also arise from invagination of the
cytoplasmic membrane, but instead of forming
vesicles, they become arranged as membrane
stacks, similar to the thylakoids of cyanobacteria
Thylakoid membranes in the cytoplasm of
cyanobacteria. Chloroplasts would be
somewhat analogous to cyanobactria being present
in the cytoplasm of algae and plant cells
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Photo-phosphorylation
Is similar to electron transport in that 1. a
proton gradient is generated to provide PMF for
ATP synthesis 2. it involves a series of
membrane bound electron acceptors (known
collectively as photosystems) 3. the membranes
involved in photosynthesis contain an ATPase
that is responsible for ATP synthesis when a
proton gradient is established Is different in
that 1. it produces reduced cofactors in the
form of NADPH 2. H2O is split into O2 to
provide electrons and protons.
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Oxidative Photophosphorylation

• Synopsis
• Carbon dioxide (to be used in Carbon fixation)
  enters the outer and inner membranes
• of the chloroplast or Cyanobacteria.
• For the Calvin cycle, CO2 is fixed in the stroma
  of the chloroplast or the cytoplasm
• of cyanobacteria
• 3. Oxidative photophosphorylation occurs in the
  thylakoid membranes that are present
• A. The photosystems and electron carriers are
  present in these membranes
• 4. H20 is split into O2 inside the thylakoid
  space of thylakoids.
• 5. Oxygen is released during oxidative
  photophosporylation and exits through the inner
• and outer membranes of the chloroplasts/cyanobacte
  ria
• NADPH and ATP is released into the stroma of
  chloroplasts/cytoplasm of
• cyanobacteria and used in CO2 fixation
• 7. Chemiosmotic theoryshuttling of protons to
  produce the PMF required for ATP
• synthesis.
• A. During the transfer of electrons through the
  e- carriers in the thylakoid
• membrane a proton gradient is achieved.
• B. Protons are pumped into the thylakoid space
  during electron transfer
• C. An ATP synthase complex is embedded in the
  thylakoid membrane
• D. When the protons flow through the ATP
  synthase complex from the thylakoid

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Enlarged thylakoid in a chloroplast or in
Cyanobacteria
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PhotosynthesisLight dependent reactions vs.
Light independent reactions (Dark Reactions)
Summary Rxn 6CO2 12H2O ? C6H12O6 6O2
6H2O OR 6CO2 6H2O ? C6H12O6 6O2
chlorophyll
Light reaction Water ADP Pi NADP
Oxygen ATP
NADPH
light
Dark reaction Carbon dioxide ATP NADPH
Glucose
enzymes
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The Z pathway is used by the aerobic
micro-organisms 1.algae 2. cyanobacteria.
Oxidative phosphorylation involved 2 photosystems
that require two separate photo absorption
acts. 1. Photosystem I (PSI)?Chlorophyll
reaction center absorbs light at 700 nm 2.
Photosystem II (PSII)?Chlorophyll reaction center
absorbs light at 680 nm
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The Z pathway allows the noncyclic flow of
electrons seen in oxidative photophosphorylation

• 1. ATP production
• 2. NADP reduced to
• NADPH for biosynthesis
• Electrons/protons from
• splitting water or exciting
• photo reaction centers

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Step by step actions in noncyclic oxidative
photo-phosphorylation

• Chlorophyll Rxn center of PSII absorbs light at
  680 nm
• The chlorophyll Rxn center becomes energetically
  excited and loses
• an electron (e-)
• The e- is transferred through a series of
  membrane bound e- carriers
• until the e- is transferred to the chlorophyll
  Rxn center of PSI
• The transfer of e-s from the Rxn center of PSII
  to PSI creates a proton
• gradient such that ATP is generated via
  chemiosmosis
• Light excites the chlorophyll Rxn center of PSI
  such that an e- is
• released from that Rxn center
• 6. e-s are transferred through a series of
  membrane bound e- carriers in
• PSI
• The protons generated through this e- transfer is
  used to reduce NADP
• to NADPH (NADPH is used in biosynthetic pathways)
• The Rxn center of PSII meanwhile has lost an e-
  that must be replaced
• This is accomplished when H2O is split to form O2
  and protons.
• The e-s are transferred thru membrane bound
  carriers to the Rxn center
• Of PSII. This also generates PMF for ATP
  synthesis.

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Anaerobic Photosynthesis (cyclic) is used by the
halophilic purple bacteria, the Heliobacteria and
the Green sulfur bacteria.
Only one photosystem is employed (PSI) The Rxn
center of PSI absorbs light at 840 nm NADPH is
usually not produced The bacteria get reducing
power from other sources (external or internal)
besides water
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Cyclic Anaerobic photo-phosphorylation

• ATP production 2. In some cases NADP reduced to
  NADPH by reverse
• electron flow 3. Reducing equivalents for
  biosynthesis can also come
• from external sources besides water or from FeS
  centers within membranes.

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Step by step actions in cyclic anaerobic
photo-phosphorylation

• Light is absorbed by the Rxn center in PSI
• e-s are transferred through membrane bound
  carriers to generate
• PMF.
• The excited e- is returned to the Rxn center of
  PSI
• Where do the bacteria get the reducing
  equivalents for biosynthesis?
• Purple bacteria reverse e- flow through the
  membrane requires
• ATP but can be used to reduce NADP to NADPH
• Green Sulfur bacteria/Heliobacteria FeS of the
  PSI can transfer
• e-s and protons to molecules to be reduced (i.e.
  CO2)
• Purple bacteria/Green sulfur bacteria Sulfide
  in the form of H2S
• or S2O3 serve as proton/electron donors that
  enter the chain at
• cytochrome c2 for extra reducing power
• (N.B. H2S or S2O3 are oxidized to elemental
  sulfur that can be stored in
• bacteria or expelled)