Methanogenesis Flashcards
What is methane?
CH4 is a waste product from metabolism
What are the 2 pathways for methanogenesis?
hydryogenotrophic
acetoclastic
What are the 3 major enzymes involved in methanogenesis?
methyl co-M reductase
formylmethanofuran dehydrogenase (hydrogentrophic)
carbon monoxide dehydrogenase (acetoclastic)
What are the major genes that code for the enzymes in the pathways?
16s rRNA (to detect bacteria or archaea)
mcrA (MCR)
fmdB (MF)
codhA (COD)
What are the 3 major types of organisms that are methanogens? What domain are they from?
Methanococcales
Methanobacteriales
Methanomicrobiales
All Archaea
T or F: some methanogens are bacteria
false, they’re all Archaea
What phylum do all the methanogens belong to?
Euryarcheota
Where do methanogens exist?
anaerobic environments
wide distribution
- sediment
- rice paddies
- guts (termites, ruminants)
What is methanogenesis?
the process of producing methane by Euryarchaeota
Under what conditions is methane formed?
only anoxic
What is the hydrogenotrophic pathway?
1 of 2 major methanogenesis pathways
the reduction of carbonates with H2 to produce
4H2 + CO2 –> CH4 + 2 H2O
What is the acetotrophic pathway?
1 of 2 major methanogenesis pathways
the fermentation of acetate
CH3COO- + H+ –> CH4 + CO2
What is the third pathway for methanogenesis that involves methanol?
methanol (CH3OH) and methylamines (CH3NH3+) can be used as substrates by some methanogens
Which pathway of methanogenesis is most common in natural environments like rice paddies or sediment?
fermentation of acetate accounts for 75-80% of methane production
T or F: methanogenesis is linked to ATP synthesis
true
Where do the precursors for methanogenesis come from?
anaerobic degradation of OM
what is a byproduct of methanogenesis?
CO2
What nutrients must be absent in order for methanogens to be active?
O2, NO3- and SO4-
Which cofactors/coenzymes are required for energy conservation?
Coenzyme F420 and MPH (Methanophenazine)
What does coenzyme F420 do?
it transfers H+ to CO2 to reduce it to CH4
What does Methanophenazine do?
accepts H+ from F42O and releases it into the periplasmic space
Which cytochrome is present in methanogens?
cytochrome b
Which type of methanogenesis would occur within the guts of termites that live in damp wood on vancouver island? why?
hydrogenotrophic because the methanogen species contain cytochrome F420
Which enzyme cofactors are unique to methanogens that use the hydrogenotrophic pathway?
Methanofuran (MF)
Methanopterin (MP)
Coenzyme F430 (related to coenzyme M)
Coenzyme B (CoB)
Which enzyme cofactor is common to both methanogenesis pathway
Coenzyme M (CoM)
Which element is core to the CoM?
Sulfur
Coenzyme F430 has which element in its core?
Ni+
Which gene is a good indicator for present and active methanogens?
mcrA genes
methyl coenzyme M reductase
What are. the 3 steps of hydrogenotrophic pathway?
- binding of CO2 to formyl by MF-dehydrogenase
- binding of H2 to formyl to reduce to methylene and then again to methyl by MP-hydrogenase reacting with F420
- binding/transfer of CH3 by CoM-reductase - reduction of methyl to methane
Which enzymes are involved in the 3 major steps of the hydrogenotrophic pathway?
- MF-dehydrogenase binds CO2 to formyl
- MP-hydrogenase reduces formyl and methylene
- CoM-reductase reduces methyl to methane
T or F: the hydrogenotrophic pathway is linked to the proton motive force and ATP production
true
which gene codes for the enzyme that binds CO2 to formyl and initiates the hydrogenotrophic pathway?
fmd
Which gene is unique to acetotrophs?
codhA
What does the codhA gene tell us about the type of methanogen present?
it’s using acetate
= acetotrophic pathway
Which gene is highly conserved within methanogens?
mcrA
What are the 3 steps in the acetotrophic pathway?
- binding of acetate to CoA and CO dehydrogenase
- transfer of CH3 from CODH to corrinoid by CO dehydrogenase
- transfer of CH3 from corrinoid to CoM by CoM-Reductase
T or F: is the acetotrophic pathway linked to ATP production
true
what is an example of a corrinoid?
vitamin b12
How does the first step of the 2 major pathways differ?
the enzymes
MF dehydrogenase for hydrogenotrophic
CO dehydrogenase for acetotrophic/acetoclastic
How does the second step of the 2 major pathways differ?
MP hydrogenase is used for hydrogenotrophic
corrinoid is used for acetotrophic/acetoclastic
How does the third step of the 2 major pathways differ?
they both use Methyl Co-M reductase
Which operon would be the target for a PCR primer?
mcr
What is QPCR?
a PCR technique that uses fluorescence with real time quantity for copies of genes
Which methanogen genomes have been sequenced?
Methanococcus jannaschii
Methanobacterium thermoautotrophicum
Methanopyrus kandleri
Methanosarcina acetivorans
Methanosarcina mazei
Describe the environment Methanococcus jannaschii is present in. Which order is it from?
named after Jannasch
Hydrogenotrophic only
thermophile
marine
What are 3 examples of methanobacteriales?
Methanobacterium thermoautotrophicum
Methanobacterium formicicum
Methanobrevibacter smithii
Describe the environment Methanobacterium thermoautotrophicum is present in. Which order is it from?
hydrogenotrophic only
thermophile
sewage sludge
freshwater (salt intolerant)
Methanobacteriales
Describe the environment Methanobacterium formicicum is present in. Which order is it from?
hydrogenotrophic only
in ruminant guts
MEthanobacteriales
Describe the environment Methanobrevibacter smithii is present in. Which order is it from?
Hydrogenotrophic only
found in the human gut
Methanobacteriales
What are 2 examples of Methanomicrobiales that can use acetate?
Methanosarcina barkeri - acetate, H+ + CO2 and methanol
Methanosaeta concilii - only acetate
Describe the environment Methanosarcina barkeri is present in. Which order is it from?
mesophile
freshwater and marine sediment
ruminant guts
can use H+ + CO2, acetate and methanol
Methanomicrobiales
Describe the environment Methanosaeta concilii is present in. Which order is it from?
acetate only
mesophile
rice paddies, sewage sludge, sanitary landfill
slow growing
Methanomicrobiale
Which example methanogen uses acetate only?
Methanosaeta concilli
How can methanogen activity be measured?
- CH4 production in soil microcosms by gas chromatography and FID or static gas chamber
- production of 14CH4 from labelled precursors to determine contribution of pathways to:
a) overall methane production
b) soil or sediment core profiles
How can methanogen abundance be measured?
- using MPN (most probable number) in liquid culture with mineral medium appropriate to growth
- 16s rRNA fluorescent probes (FISH)
- mcrA gene detection by qPCR or probing
Which gene would be used to determine the presence of methanogens?
mcrA
Which gene tells you the methanogen is using CO2 and not acetate?
fwdB or fmdB (different metalloproteins)
How can methanogen diversity be measured?
- culture-dependent methods: isolation from MPN tube with highest dilution
- culture-independent methods: cloning of amplicons, sequencing, sequence analysis with database
How did the presence of a nitrate + acetate mixture and acetate influence production of methane in rice paddies in Po Valley when considering interactions between methanogens and denitrifying bacteria?
presence of nitrate suppressed methane production
What does it mean that the Po Valley study found that there was no competition for substrates, but the methanogens were suppressed?
denitrifying bacteria and methanogens are not using the same source, but may be competing for an intermediate substrate
What implications does understanding how to suppress methane production have?
control of methane emissions requires an understadnign for suppressing methanogens - could be with addition of N in fertilizers or aeration (methanogens can’t function in oxic conditions)
T or F: Archaea are not susceptible to antibiotics - why/why not?
true, unlike bacteria, they do not have peptidoglycan in their cell walls
How can antibiotics be used to isolate methanogens?
methanogens do not have peptidoglycan in their cell walls like bacteria do, the addition of antibiotics to their culture will kill off the bacteria present and allow Archaea to survive
What environmental conditions limit methanogenesis?
presence of oxygen is toxic to methanogens
availability of acetate and hydrogen
presence of oxidants (suppressors) like NO3-, SO42-, Fe3+, Mn4+
- competitive mechanism
- amensalism (produces toxic intermediate)
Which substrates suppress methanogens?
oxidants like NO3-, SO42-, Fe3+, Mn4+
What is amensalism?
when a toxic intermediate is produced
Where are methanogens usually found?
only anoxic environments
flooded soils, sediment or freshwater
marine sediment
microbial mats
digestive tracts or rumen
What level of contribution do methanogens have to organic matter degradation in flooded soils, sediment, or freshwater ecosystems?
large (40-60%)
due to lower sulfate concentrations
Do methanogens contribute more or less to degradation of OM in marine or freshwater environments? why?
more in freshwater because less sulfate
most of the degradation of OM in marine is from sulfate-reduing bacteria
Which methanogen habitats have highest CH4 release?
ruminant guts and rice paddies
natural wetlands
landfills
What are deep sea methane hydrates?
ice lattices that surround methane molecules located at the edges of continental plates
What causes the formation of deep sea methane hydrates?
high hydrostatic pressure and low temperatures (very deep in ocean)
how could deep sea methane hydrates be exploited for energy?
they are huge resources of methane but the release of them would emit a lot of CO2
Where are some deep sea methane hydrates located?
Nankai Through
Eel River
Cascadia Mountains
Gulf of Mexico
Barkley Canyon
What is acetogenesis?
the production of CO2 from acetate by acetogens
What is the net reaction for acetogenesis?
4 H2 + H+ + 2 HCO3- = CH3COO- + 4 H2O
Does methanogenesis produce more or less energy than acetogenesis?
more energy
In what conditions do acetogens dominate over methanogens in competition for CO2 + H2?
in low temperatures, otherwise methanogens outcompete
