Methanogen / Methanotroph

From Wikipedia, the free encyclopedia

Methanogen

Methanogens are microorganisms that produce methane as a metabolic byproduct in hypoxic conditions. They are prokaryotic and belong to the domain of archaea. They are common in wetlands, where they are responsible for marsh gas, and in the digestive tracts of animals such as ruminants and humans, where they are responsible for the methane content of belching in ruminants and flatulence in humans. In marine sediments the biological production of methane, also termed methanogenesis, is generally confined to where sulfates are depleted, below the top layers. Moreover, methanogenic archaea populations play an indispensable role in anaerobic wastewater treatments. Others are extremophiles, found in environments such as hot springs and submarine hydrothermal vents as well as in the "solid" rock of Earth's crust, kilometers below the surface.

Physical description

Methanogens are coccoid (spherical shaped) or bacilli (rod shaped). There are over 50 described species of methanogens, which do not form a monophyletic group, although all known methanogens belong to Archaea. They are mostly anaerobic organisms that cannot function under aerobic conditions, but recently a species (Candidatus Methanothrix paradoxum) has been identified that can function in anoxic microsites within aerobic environments. They are very sensitive to the presence of oxygen even at trace level. Usually, they cannot sustain oxygen stress for a prolonged time. However, Methanosarcina barkeri is exceptional in possessing a superoxide dismutase (SOD) enzyme, and may survive longer than the others in the presence of O₂. Some methanogens, called hydrogenotrophic, use carbon dioxide (CO₂) as a source of carbon, and hydrogen as a reducing agent.

The reduction of carbon dioxide into methane in the presence of hydrogen can be expressed as follows:

CO₂ + 4 H₂ → CH₄ + 2H₂O

Some of the CO₂ reacts with the hydrogen to produce methane, which creates an electrochemical gradient across the cell membrane, used to generate ATP through chemiosmosis. In contrast, plants and algae use water as their reducing agent.

Methanogens lack peptidoglycan, a polymer that is found in the cell walls of Bacteria but not in those of Archaea. Some methanogens have a cell wall that is composed of pseudopeptidoglycan. Other methanogens do not, but have at least one paracrystalline array (S-layer) made up of proteins that fit together like a jigsaw puzzle.

Extreme living areas

Methanogens play a vital ecological role in anaerobic environments of removing excess hydrogen and fermentation products that have been produced by other forms of anaerobic respiration. Methanogens typically thrive in environments in which all electron acceptors other than CO₂ (such as oxygen, nitrate, ferriciron (Fe(III)), and sulfate) have been depleted. In deep basaltic rocks near the mid ocean ridges, they can obtain their hydrogen from the serpentinisation reaction of olivine as observed in the hydrothermal field of Lost City. 

The thermal breakdown of water and water radiolysis are other possible sources of hydrogen. 

Methanogens are key agents of remineralization of organic carbon in continental margin sediments and other aquatic sediments with high rates of sedimentation and high sediment organic matter. Under the correct conditions of pressure and temperature, biogenic methane can accumulate in massive deposits of methane clathrates, which account for a significant fraction of organic carbon in continental margin sediments and represent a key reservoir of a potent greenhouse gas.

Methanogens have been found in several extreme environments on Earth – buried under kilometres of ice in Greenland and living in hot, dry desert soil. They are known to be the most common archaebacteria in deep subterranean habitats. Live microbes making methane were found in a glacial ice core sample retrieved from about three kilometres under Greenland by researchers from the University of California, Berkeley. They also found a constant metabolism able to repair macromolecular damage, at temperatures of 145 to –40 °C.

Another study has also discovered methanogens in a harsh environment on Earth. Researchers studied dozens of soil and vapour samples from five different desert environments in Utah, Idaho and California in the United States, and in Canada and Chile. Of these, five soil samples and three vapour samples from the vicinity of the Mars Desert Research Station in Utah were found to have signs of viable methanogens.

Some scientists have proposed that the presence of methane in the Martian atmosphere may be indicative of native methanogens on that planet.

Closely related to the methanogens are the anaerobic methane oxidizers, which utilize methane as a substrate in conjunction with the reduction of sulfate and nitrate. Most methanogens are autotrophic producers, but those that oxidize CH₃COO⁻ are classed as chemotroph instead.

Comparative genomics and molecular signatures

Comparative genomic analysis has led to the identification of 31 signature proteins which are specific for methanogens (also known as Methanoarchaeota). Most of these proteins are related to methanogenesis, and they could serve as potential molecular markers for methanogens. Additionally, 10 proteins found in all methanogens which are shared by Archaeoglobus, suggest that these two groups are related. In phylogenetic trees, methanogens are not monophyletic and they are generally split into three clades. Hence, the unique shared presence of large numbers of proteins by all methanogens could be due to lateral gene transfers.

Metabolism

Methane production

Methanogens are known to produce methane from substrates such as H₂/CO₂, acetate, formate, methanol and methylamines in a process called methanogenesis. Different methanogenic reactions are catalyzed by unique sets of enzymes and coenzymes. While reaction mechanism and energetics vary between one reaction and another, all of these reactions contribute to net positive energy production by creating ion concentration gradients that are used to drive ATP synthesis. The overall reaction for H₂/CO₂ methanogenesis is:

${\displaystyle {\ce {CO2 + 4 H2 -> CH4 + 2 H2O}}}$
  (∆G˚’ = -134 kJ/mol CH₄)

Well-studied organisms that produce methane via H2/CO₂ methanogenesis include Methanosarcina barkeri, Methanobacterium thermoautotrophicum, and Methanobacterium wolfei. These organism are typically found in anaerobic environments.

In the earliest stage of H₂/CO₂ methanogenesis, CO₂ binds to methylfuran (MF) and is reduced to formyl-MF. This endergonic reductive process (∆G˚’= +16 kJ/mol) is dependent on the availability of H₂ and is catalyzed by the enzyme formyl-MF dehydrogenase.

${\displaystyle {\ce {CO2 + H2 + MF -> HCO-MF + H2O}}}$

The formyl constituent of formyl-MF is then transferred to the coenzyme tetrahydromethanopterin (H4MPT) and is catalyzed by a soluble enzyme known as formyl transferase. This results in the formation of formyl-H4MPT.

${\displaystyle {\ce {HCO-MF + H4MPT -> HCO-H4MPT + MF}}}$

Formyl-H4MPT is subsequently reduced to methenyl-H4MPT. Methenyl-H4MPT then undergoes a one-step hydrolysis followed by a two-step reduction to methyl-H4MPT. The two-step reversible reduction is assisted by coenzyme F₄₂₀ whose hydride acceptor spontaneously oxidizes. Once oxidized, F₄₂₀’s electron supply is replenished by accepting electrons from H₂. This step is catalyzed by methylene H4MPT dehydrogenase.

${\displaystyle {\ce {HCO-H4MPT + H+ -> CH-H4MPT+ + H2O}}}$ 

(Formyl-H4MPT reduction)

${\displaystyle {\ce {CH-H4MPT+ + F420H2 -> CH2=H4MPT + F420 + H+}}}$

 

(Methenyl-H4MPT hydrolysis)

${\displaystyle {\ce {CH2=H4MPT + H2 -> CH3-H4MPT + H+}}}$ 

(H4MPT reduction)

Next, the methyl group of methyl-M4MPT is transferred to coenzyme M via a methyltransferase-catalyzed reaction.

${\displaystyle {\ce {CH3-H4MPT + HS-CoM -> CH3-S-CoM + H4MPT}}}$

The final step of H₂/CO₂ methanogenic involves methyl-coenzyme M reductase and two coenzymes: N-7 mercaptoheptanoylthreonine phosphate (HS-HTP) and coenzyme F₄₃₀. HS-HTP donates electrons to methyl-coenzyme M allowing the formation of methane and mixed disulfide of HS-CoM.^[24] F₄₃₀, on the other hand, serves as a prosthetic group to the reductase. H₂ donates electrons to the mixed disulfide of HS-CoM and regenerates coenzyme M.

${\displaystyle {\ce {CH3-S-CoM + HS-HTP -> CH4 + CoM-S-S-HTP}}}$ 

  (Formation of methane)

${\displaystyle {\ce {CoM-S-S-HTP + H2 -> HS-CoM + HS-HTP}}}$ 

(Regeneration of coenzyme M)

Wastewater treatment

Methanogens are widely used in anaerobic digestors to treat wastewater as well as aqueous organic pollutants. Industries have selected methanogens for their ability to perform biomethanation during wastewater decomposition thereby rendering the process sustainable and cost-effective.

Bio-decomposition in the anaerobic digester involves a four-staged cooperative action performed by different microorganisms. The first stage is the hydrolysis of insoluble polymerized organic matter by anaerobes such as Streptococcus and Enterobacterium. In the second stage, acidogens breakdown dissolved organic pollutants in wastewater to fatty acids. In the third stage, acetogens convert fatty acids to acetates. In the final stage, methanogens metabolize acetates to gaseous methane. The byproduct methane leaves the aqueous layer and serves as an energy source to power wastewater-processing within the digestor, thus generating a self-sustaining mechanism.

Methanogens also effectively decrease the concentration of organic matter in wastewater run-off. For instance, agricultural wastewater, highly rich in organic material, has been a major cause of aquatic ecosystem degradation. The chemical imbalances can lead to severe ramifications such as eutrophication. Through anaerobic digestion, the purification of wastewater can prevent unexpected blooms in water systems as well as trap methanogenesis within digesters. This allocates biomethane for energy production and prevents a potent greenhouse gas, methane, from being released into the atmosphere. 

The organic components of wastewater vary vastly. Chemical structures of the organic matter select for specific methanogens to perform anaerobic digestion. An example is the members of Methanosaeta genus dominate the digestion of palm oil mill effluent (POME) and brewery waste. Modernizing wastewater treatment systems to incorporate higher diversity of microorganisms to decrease organic content in treatment is under active research in the field of microbiological and chemical engineering. Current new generations of Staged Multi-Phase Anaerobic reactors and Upflow Sludge Bed reactor systems are designed to have innovated features to counter high loading wastewater input, extreme temperature conditions, and possible inhibitory compounds.

Methanotroph

Methanotrophs (sometimes called methanophiles) are prokaryotes that metabolize methane as their only source of carbon and energy. They can be either bacteria or archaea and can grow aerobically or anaerobically, and require single-carbon compounds to survive.

General

Methanotrophs are especially common in or near environments where methane is produced, although also methanotrophs exist that can oxidize atmospheric methane. Their habitats include wetlands, soils, marshes, rice paddies, landfills, aquatic systems (lakes, oceans, streams) and more. They are of special interest to researchers studying global warming, as they play a significant role in the global methane budget, by reducing the amount of methane emitted to the atmosphere.

Methanotrophy is a special case of methylotrophy, using single-carbon compounds that are more reduced than carbon dioxide. Some methylotrophs, however, can also make use of multi-carbon compounds which differentiates them from methanotrophs that are usually fastidious methane and methanol oxidizers. The only facultative methanotrophs isolated to date are members of the genus Methylocella and Methylocystis.

In functional terms, methanotrophs are referred to as methane-oxidizing bacteria, however, methane-oxidizing bacteria encompass other organisms that are not regarded as sole methanotrophs. For this reason methane-oxidizing bacteria have been separated into four subgroups: two methane-assimilating bacteria (MAB) groups, the methanotrophs, and two autotrophic ammonia-oxidizing bacteria (AAOB).

Methanotroph classification

Methantrophs can be either bacteria or archaea. Which methanotroph species is present, is mainly determined by the availability of electron acceptors. Many types of methane oxidizing bacteria (MOB) are known. Differences in the method of formaldehyde fixation and membrane structure divide these bacterial methanotrophs into several groups. These include the Methylococcaceae and Methylocystaceae. Although both are included among the Proteobacteria, they are members of different subclasses. Other methanotroph species are found in the Verrucomicrobiae. Among the methanotrophic archaea, several subgroups are determined.

Aerobic methanotrophs

Under aerobic conditions, methanotrophs combine oxygen and methane to form formaldehyde, which is then incorporated into organic compounds via the serine pathway or the ribulose monophosphate (RuMP) pathway, and [Carbon dioxide], which is released. Type I and type X methanotrophs are part of the Gammaproteobacteria and they use the RuMP pathway to assimilate carbon. Type II methanotrophs are part of the Alphaproteobacteria and utilize the serine pathway of carbon assimilation. They also characteristically have a system of internal membranes within which methane oxidation occurs. No methanotrophic archaea are capable of using oxygen.

Anaerobic methanotrophs

Under anoxic conditions, methanotrophs use different electron acceptors for methane oxidation. This can happen in anoxic habitats such as marine or lake sediments, oxygen minimum zones, anoxic water columns, rice paddies and soils. Some specific methanotrophs can reduce nitrate or nitrite, and couple that to methane oxidation. Investigations in marine environments revealed that methane can be oxidized anaerobically by consortia of methane oxidizing archaea and sulfate-reducing bacteria. This type of Anaerobic oxidation of methane (AOM) mainly occurs in anoxic marine sediments. The exact mechanism behind this is still a topic of debate but the most widely accepted theory is that the archaea use the reversed methanogenesis pathway to produce carbon dioxide and another, unknown substance. This unknown intermediate is then used by the sulfate-reducing bacteria to gain energy from the reduction of sulfate to hydrogen sulfide. The anaerobic methanotrophs are not related to the known aerobic methanotrophs; the closest cultured relative to the anaerobic methanotrophs are the methanogens in the order Methanosarcinales. Metal-oxides, such as manganese and iron, can also be used as terminal electron acceptors by ANME. For this, no consortium is needed. ANME shuttle electrons directly to the abiotic particles, which get reduced chemically. 

In some cases, aerobic methane oxidation can take place in anoxic (no oxygen) environments. Candidatus Methylomirabilis oxyfera belongs to the phylum NC10 bacteria, and can catalyze nitrite reduction through an “intra-aerobic” pathway, in which internally produced oxygen is used to oxidise methane. In clear water lakes, methanotrophs can live in the anoxic water column, but receive oxygen from photosynthetic organisms, that they then directly consume to oxidise methane aerobically.

Special methanotroph species

Methylococcus capsulatus is utilised to produce animal feed from natural gas.

Recently, a new bacterium Candidatus Methylomirabilis oxyfera was identified that can couple the anaerobic oxidation of methane to nitrite reduction without the need for a syntrophic partner. Based on the studies of Ettwig et al., it is believed that M. oxyfera oxidizes methane anaerobically by utilizing the oxygen produced internally from the dismutation of nitric oxide into nitrogen and oxygen gas.

Properties

RuMP pathway in type I methanotrophs

 

Serine pathway in type II methanotrophs

 

Methanotrophs oxidize methane by first initiating reduction of an oxygen atom to H₂O₂ and transformation of methane to CH₃OH using methane monooxygenases (MMOs). Furthermore, two types of MMO have been isolated from methanotrophs: soluble methane monooxygenase (sMMO) and particulate methane monooxygenase (pMMO). Cells containing pMMO have demonstrated higher growth capabilities and higher affinity for methane than sMMO containing cells. It is suspected that copper ions may play a key role in both pMMO regulation and the enzyme catalysis, thus limiting pMMO cells to more copper-rich environments than sMMO producing cells.