Microbial C1-Fixation
Dr. Simon K.-M. R. Rittmann
Ferry, 2010
Global carbon cycle
The global carbon cycle.
Aerobic O2-requiring
conversions are shown in the
top panel and anaerobic
conversions in the bottom
panel. Step 1: Fixation of CO2
into organic matter. Step 2:
Decomposition of organic
matter to CO2 by O2-requiring
microbes. Step 3: Deposition
of organic matter into
anaerobic environments. Step
4: Decomposition of complex
biomass by fermentative
microbes. Step 5: Conversion
of volatile fatty acids by
obligate H2- and formateproducing
syntrophic
acetogens. Step 6: H2- and
formate-dependent reduction of
CO2 to CH4 and conversion of
the methyl group of acetate to
CH4 by methanogens. Step 7:
Anaerobic oxidation of CH4.
Step 8: Diffusion of CH4 into
aerobic zones. Step 9: Aerobic
oxidation of CH4 by O2requiring
methylotrophs.
2
(Dissolved) carbon dioxide (CO2),
also in the form of bicarbonate (HCO3
–)
Carbon monoxide (CO)
Formate (HCOO–)
Methane (CH4)
Methanol (CH3OH)
Mono-, di-, trimethylamine
Methanethiole (CH3SH)
C1 species
3
CO2 equilibrium in aqueous solution
Wang et al., 2010
4
Currently six CO2 fixation pathways are known:
Calvin–Benson–Bassham cycle
reductive tricarboxylic acid cycle
reductive acetyl CoA pathway – Wood-Ljungdahl pathway
3-hydroxypropionate bicycle
3 hydroxypropionate/4 hydroxybutyrate cycle
dicarboxylate/4 hydroxybutyrate cycle
(reductive hexulose-phosphate pathway) proposed
[reductive glycine pathway] metagenome
[Crotonyl-CoA/ethylmalonyl-CoA/hydroxybutyryl-CoA cycle]
synthetic
CO2 fixation pathways
5
 In the Calvin–Benson–Bassham cycle, which was discovered about 50 years
ago, CO2 reacts with the five-carbon sugar ribulose 1,5-bisphosphate to yield two
carboxylic acids, 3-phosphoglycerate, from which the sugar is regenerated.
 This cycle operates in plants, algae, cyanobacteria, some aerobic or facultative
anaerobic Proteobacteria, CO-oxidizing mycobacteria and representatives of the
genera Sulfobacillus (iron- and sulphur-oxidizing Firmicutes) and Oscillochloris
(green sulphur bacteria).
 An autotrophic symbiotic cyanobacterium conferred the CO2 fixation machinery
on a eukaryotic cell giving rise to the chloroplasts of plant cells. The presence of
the key enzyme, ribulose 1,5-bisphosphate carboxylase–oxygenase
(RubisCO), is often considered to be synonymous with autotrophy.
 Phylogenetic analysis and general considerations denote the Calvin cycle as a
late innovation.
Calvin–Benson–Bassham cycle
Berg et al., 2010
6
Diagram illustrating the reactions of the Calvin–Benson–Bassham cycle. The corresponding enzymes are: 1: ribulose-1,5-bisphosphate carboxylase/oxygenase, 2: phosphoglycerate
kinase, 3: glyceraldehyde-3-phosphate dehydrogenase, 4: triose phosphate isomerase, 5: fructose-bisphosphate aldolase, 6: fructose-1,6-bisphosphatase, 7: sedoheptulose bisphosphatase, 8:
transketolase, 9: ribose-5-phosphate isomerase, 10: ribulose-5-phosphate 3-epimerase and 11: phosphoribulokinase. The directions of the arrows represent the direction of the pathway, and do
not indicate that the enzyme reactions are irreversible.
Calvin–Benson–Bassham cycle
Sato & Atomi , 2010
7
 In 1966, Arnon, Buchanan and co-workers proposed another autotrophic cycle for
the green sulphur bacterium Chlorobium limicola, the reductive citric acid cycle
(also known as the Arnon–Buchanan cycle).
 This cycle is less energy-consuming than the Calvin cycle, involves enzymes that
are sensitive to oxygen and is therefore found only in anaerobes or in aerobes
growing at low oxygen tensions.
 These include some Proteobacteria, green sulphur bacteria and microaerophilic
bacteria of the early bacterial phylum Aquificae.
 Initially, the reductive citric acid cycle was also proposed to operate in certain
archaea (notably Thermoproteus neutrophilus), but recent findings refute this
proposal.
Berg et al., 2010
reductive tricarboxylic acid cycle
8
Diagram illustrating the reactions of the reductive
tricarboxylic acid cycle. The corresponding enzymes are:
1: malate dehydrogenase,
2: fumarate hydratase,
3: fumarate reductase,
4: succinyl-CoA synthetase (acetyl-CoA: succinate CoA
transferase is used in Desulfobacter hydrogenophilus),
5: oxoglutarate synthase,
6: isocitrate dehydrogenase,
7: aconitate hydratase,
8: ATP-citrate lyase,
9: pyruvate synthase,
10: pyruvate carboxylase,
11: phosphoenolpyruvate synthase and
12: phosphoenolpyruvate carboxylase.
The directions of the arrows represent the direction of the
pathway, and do not indicate that the enzyme reactions are
irreversible.
reductive tricarboxylic acid cycle
Sato & Atomi , 2010
9
 At the start of the 1980s, a third autotrophic pathway was found in certain Grampositive
bacteria and methane-forming archaea, the reductive acetyl-coenzyme A
(acetyl-CoA) or Wood–Ljungdahl pathway.
 In these strict anaerobic organisms that now also include some Proteobacteria,
Planctomycetes, spirochaetes and Euryarchaeota, one CO2 molecule is reduced
to CO and one to a methyl group (bound to a carrier); subsequently, acetyl-CoA
is synthesized from CO and the methyl group.
 Although this pathway is the most energetically favourable autotrophic carbon
fixation pathway, it is restricted to strictly anaerobic organisms.
 Use of the acetyl-CoA pathway as an autotrophic, terminal electron-accepting
process is the hallmark of acetogens. Other obligate anaerobic bacteriological
groups, including methanogens and sulfate-reducing bacteria, also utilize the
acetyl-CoA pathway for either catabolic or anabolic purposes.
Berg et al., 2010; Drake et al., 1997
reductive acetyl CoA pathway
10
During the reductive synthesis of acetate via the acetyl-CoA pathway,
six reducing equivalents are required for the fixation of CO2 to the
methyl level, and two reducing equivalents are required for the fixation
of CO2 to the carbonyl level.
The overall conversion of substrate to product can be divided into
oxidative and reductive processes. For example, the glycolytic
utilization of glucose yields eight reducing equivalents per glucose
consumed that are routed towards the reductive synthesis of acetate.
reductive acetyl CoA pathway
Drake et al., 1997
11
Drake et al., 2008
Homoacetogenic conversion of
glucose to acetate.
The two molecules of CO2 that are
reduced to acetate in the acetyl-CoA
pathway can be derived from
exogenous CO2 rather than the CO2
that is produced via the
decarboxylation of pyruvate (see
text). ABBREVIATIONS: ATPSLP =
ATP that is produced by substratelevel
phosphorylation; [e−], reducing
equivalent.
reductive acetyl CoA pathway
12
reductive acetyl CoA pathway
Schuchmann & Müller, 2014
13
14
Excursion: flavin-based electron bifuraction
reductive acetyl CoA pathway
Schuchmann & Müller, 2016
15
16
reductive acetyl CoA pathway
Schuchmann & Müller, 2014
reductive acetyl CoA pathway
Schuchmann & Müller, 2016
17
reductive acetyl CoA pathway
18
19
reductive acetyl CoA pathway
Schuchmann & Müller, 2014
20
reductive acetyl CoA pathway
Schuchmann & Müller, 2014
reductive acetyl CoA pathway
Hypothetical scheme for the
differential metabolic capacities of
the acetogen Peptostreptococcus
productus U-1. Abbreviations: LDH:
lactate dehydrogenase; FBP: fructose-
1,6-bisphosphate; DHAP:
dihydroxyacetone phosphate; GAP:
glyceraldehyde-3-phosphate; OAA:
oxaloacetate. Broken line indicates
stimulation of lactacte dehydrogenase
by fructose- 1,6-bisphosphate.
Drake et al., 1997
21
reductive acetyl CoA pathway
Drake et al., 1997, 2008
22
Representative oxidizeable substrates that can be used for the reductive synthesis of acetate via the acetyl-CoA ‘Wood/Ljungdahl’ pathway
Drake et al., 1997
reductive acetyl CoA pathway
23
reductive acetyl CoA pathway
Diagram illustrating the reactions of the reductive acetyl-CoA pathway.
Coloured pathways indicate related enzymes involved in methanogenesis and
acetogenesis. The corresponding enzymes are:
1: formate dehydrogenase,
2: formyl-H4FA-synthase,
3: methenyl-H4FA-cyclohydrolase,
4: methylene-H4FA-dehydrogenase,
5: methylene-H4FA-reductase,
6: methyltransferase,
7: carbon monooxide dehydrogenase/acetyl-CoA synthase,
8: phosphotransacetylase,
9: acetate kinase,
10: formylMF dehydrogenase,
11: formylMF:H4MPT formyltransferase,
12: N5, N10-methenyl-H4MPT cyclohydrolase,
13: N5, N10-methylene-H4MPT dehydrogenase,
14: N5, N10-methylene-H4MPT reductase,
15: N5-methyl-H4MPT:coenzyme M methyltransferase,
16: methyl-coenzyme M reductase and
17: heterodisulfide reductase.
The directions of the arrows represent the direction of the pathway, and do not
indicate that the enzyme reactions are irreversible. Abbreviations are: H4FA,
tetrahydrofolate; MF, methanofuran; H4MPT, tetrahydromethanopterin; CoFe/S-P,
corrinoid-iron sulfur protein.
Sato & Atomi , 2010
24
Metabolic processes by which carbon
monoxide is utilized by Methanosarcina
acetivorans.
(A) Oxidation of carbon monoxide;
(B) reductive synthesis of acetate via the acetylCoA
pathway;
(C) production of methane.
Parenthetical numbers identify enzymes that
catalyze the indicated reactions:
1, CO dehydrogenase/acetyl-CoA synthetase;
2, formyl-methanofuran dehydrogenase;
3, formyl-methanofuran:H4MPT
formyltransferase;
4, combined activities of methenyl-H4MPT
cyclohydrolase, methylene-H4MPT
dehydrogenase; methylene-H4MPT reductase;
5, phosphotransacetylase;
6, acetate kinase;
7, methyl-H4MPT:CoM methyltransferase;
8, combined activities of methyl-CoM reductase,
membranous heterodisulfide reductase, and
membranous F420H2 dehydrogenase complex
(F420, coenzyme F420).
ABBREVIATIONS: MF = methanofuran; H4MPT
= tetrahydromethanopterin; HSCoM = coenzyme
M. [The figure is based on information in Lessner
et al. (further details on how these processes are
coupled to the chemiosmotic conservation of
energy can be found in this reference).]
reductive acetyl-coenzyme A pathway
25
reductive acetyl-coenzyme A pathway
Goyal et al., 2014
26
(1) Formylmethanofuran dehydrogenase (Fmd); (2) Formylmethanofuran:tetramethanopterin formyltransferase (Ftr), (3) methenyl-tetrahydromethanopterin cyclohydrolase (Mch),
(4) methylene-tetrahydromethanopterin dehydrogenase (Hmd), (5) methylene-tetrahydromethanopterin reductase (Mer), (6) CO dehydrogenase-acetyl-CoA synthase
Berg et al., 2010
reductive acetyl CoA pathway
27
reductive acetyl-coenzyme A pathway
Metabolic pathway map
of genes encoded in the
SM1 euryarchaeal
pangenome. Membrane
proteins are indicated by
symbols or protein
structures of subunits
(taken from Pfam
database). Predicted
proteins are indicated by
numbers, which can be
found in Supplementary
Table 2. Grey numbers
were found in SM1-MSI but
not for SM1-CG. Specific
enzymes were tested for
presence of corresponding
mRNA: Enzymes labeled in
green were also detected
in the mRNA pool of the
biofilm, while enzymes with
red color were absent in
mRNA pool. Interrogation
marks indicate potential
metabolic pathway
reactions that are likely
present but are lacking
evidence in the fragmented
genomic bins.
Probst et al., 2015
28
 The 3-hydroxypropionate bicycle (3 hydroxypropionate/malyl CoA cycle)
occurs in some green non-sulphur bacteria of the family Chloroflexaceae. This
seems to be a singular invention, and the pathway has not been found elsewhere.
 The conversion of acetyl-CoA plus two bicarbonates to succinyl-CoA uses the
same intermediates as in the hydroxypropionate–hydroxybutyrate cycle, but most
of the enzymes are completely different.
 The regeneration of acetyl-CoA proceeds by the cleavage of malyl-CoA, yielding
acetyl-CoA and glyoxylate. The assimilation of glyoxylate requires a second cycle
(hence the name bicycle).
3-hydroxypropionate bicycle
Berg et al., 2010
29
Diagram illustrating the reactions of the 3-hydroxypropionate bicycle (3-hydroxypropionate/malyl-CoA cycle). The corresponding enzymes are: 1: acetyl-CoA carboxylase, 2:
malonyl-CoA reductase, 3: propionyl-CoA synthase, 4: propionyl-CoA carboxylase, 5: methylmalonyl-CoA epimerase, 6: methylmalonyl-CoA mutase, 7:
succinyl-CoA:(S)-malate-CoA transferase, 8: succinate dehydrogenase, 9: fumarate hydratase, 10-1, 10-2, 10-3: (S)-malyl-CoA/β-methylmalyl-CoA/(S)-citramalyl-CoA
(MMC) lyase, 11: mesaconyl-C1-CoA hydratase (β-methylmalyl-CoA dehydratase), 12: mesaconyl-CoA C1-C4 CoA transferase and 13: mesaconyl-C4-CoA hydratase. The
directions of the arrows represent the direction of the pathway, and do not indicate that the enzyme reactions are irreversible.
3-hydroxypropionate bicycle
Sato & Atomi , 2010
30
Pathways of autotrophic CO2 fixation in crenarchaeota. The dicarboxylate–hydroxybutyrate cycle functions in Desulfurococcales and Thermoproteales (a) and the
hydroxypropionate–hydroxybutyrate cycle functions in Sulfolobales and Thaumarchaeota (b). Note that succinyl-coenzyme A (succinyl-CoA) reductase in Thermoproteales
and Sulfolobales uses NADPH and reduced methyl viologen (possibly as a substitute for reduced ferredoxin) in Desulfurococcales. In Sulfolobales, pyruvate might be
derived from succinyl-CoA by C4 decarboxylation. CoASH, coenzyme A; Fdred 2–, reduced ferredoxin; Fdox, oxidized ferredoxin; PEP, phosphoenolpyruvate. Berg et al., 2010
pathways of autotrophic CO2 fixation in crenarchaeota
dicarboxylate–hydroxybutyrate cycle hydroxypropionate–hydroxybutyrate cycle
31
 The 3 hydroxypropionate/4 hydroxybutyrate cycle (hydroxypropionate–
hydroxybutyrate cycle) was discovered by Berg et al. in 2007.
 The hydroxypropionate–hydroxybutyrate cycle occurs in Sulfolobales and
Thaumarchaeota. Although some of the intermediates and the carboxylation
reactions are the same as in the 3-hydroxypropionate bicycle in Chloroflexaceae,
the archaeal cycle probably has evolved independently.
Berg et al., 2007, 2010
3 hydroxypropionate/4 hydroxybutyrate cycle
32
3 hydroxypropionate/4 hydroxybutyrate cycle
Proposed autotrophic 3-hydroxypropionate/4hydroxybutyrate
cycle in M. sedula. Reactions
of the cycle are shown.
Enzymes:
1, acetyl-CoA carboxylase;
2, malonyl-CoA reductase (NADPH);
3, malonate semialdehyde reductase (NADPH);
4, 3-hydroxypropionyl-CoA synthetase (AMP-
forming);
5, 3-hydroxypropionyl-CoA dehydratase;
6, acryloyl-CoA reductase (NADPH);
7, propionyl-CoA carboxylase;
8, methylmalonyl-CoA epimerase;
9, methylmalonyl-CoA mutase;
10, succinyl-CoA reductase (NADPH);
11, succinate semialdehyde reductase (NADPH);
12, 4-hydroxybutyryl-CoA synthetase (AMP-
forming);
13, 4-hydroxybutyryl-CoA dehydratase;
14, crotonyl-CoA hydratase;
15, 3-hydroxybutyryl-CoA dehydrogenase
(NAD+);
16, acetoacetyl-CoA β-ketothiolase.
The proposed pathway of glyceraldehyde 3phosphate
synthesis from acetyl-CoA and CO2 is
also shown.
Enzymes:
17, pyruvate synthase;
18, pyruvate, water dikinase
[phosphoenolpyruvate (PEP) synthase];
19, enolase;
20, phosphoglycerate mutase;
21, 3-phosphoglycerate kinase;
22, glyceraldehyde 3-phosphate dehydrogenase.
Berg et al., 2007
33
Reactions of the crenarchaeal and thaumarchaeal variants of the HP/ HB cycle.
The reactions determining energy efficiency of the crenarchaeal (M. sedula) cycle are
shown in green, and those for the N. maritimus variant are shown in red. Reactions
common to both are shown in black. Note that although the two pathways have similar
reactions and intermediates, they are significantly different in energy efficiency and evolved
independently in Crenarchaeota and Thaumarchaeota. The numbers in square brackets
represent moles of high-energy anhydride bonds of ATP required to form 1 mol of the
corresponding central precursor metabolites.
Enzymes as numbered in circles are:
1, 3-hydroxypropionyl-CoA synthetase (ADP-forming);
2, 3-hydroxypropionyl-CoA synthetase (AMP-forming);
3, 3-hydroxypropionyl-CoA dehydratase;
4, 4-hydroxybutyryl-CoA synthetase (ADP-forming);
5, 4-hydroxybutyryl-CoA synthetase (AMP-forming);
6, 4-hydroxybutyryl-CoA dehydratase;
7, crotonyl-CoA hydratase;
8, succinyl-CoA synthetase (ADP-forming);
9, succinic semialdehyde dehydrogenase;
10, pyruvate-phosphate dikinase; and
11, pyruvatewater dikinase.
Catalytic properties of recombinant enzymes
Könnecke et al,. 2015
3 hydroxypropionate/4 hydroxybutyrate cycle
34
3 hydroxypropionate/4 hydroxybutyrate cycle
35
3 hydroxypropionate/4 hydroxybutyrate cycle
36
0,0
1000,0
2000,0
3000,0
4000,0
5000,0
6000,0
0,0 50,0 100,0 150,0 200,0 250,0
c(NO2
-)[µmolL-1]
time [h]
c(NO2-)
Results of quadruplicate batch fermentation of N. viennensis in a bioreactor system with continuous supply of 20% CO2, 21% O2, rest N2.
Fed-batch fermentation of N. viennensis
37
 The dicarboxylate/4 hydroxybutyrate cycle was discovered by Huber et al. in
2008.
 The dicarboxylate–hydroxybutyrate cycle occurs in the anaerobic crenarchaeal
orders Thermoproteales and Desulfurococcales.
Huber et al., 2008
dicarboxylate/4 hydroxybutyrate cycle
38
Diagram illustrating the reactions of the dicarboxylate/4-hydroxybutyrate cycle. The corresponding enzymes are: 1: pyruvate synthase, 2: pyruvate:water dikinase, 3:
phosphoenolpyruvate carboxylase, 4: malate dehydrogenase, 5: fumarate hydratase, 6: fumarate reductase, 7: succinate thiokinase, 8: succinyl-CoA reductase, 9: succinate
semialdehyde reductase, 10: 4-hydroxybutyryl-CoA synthetase, 11: 4-hydroxybutyryl-CoA dehydratase, 12: crotonyl-CoA hydratase, 13: 3-hydroxybutyryl-CoA
dehydrogenase and 14: acetoacetyl-CoA β-ketothiolase. The directions of the arrows represent the direction of the pathway, and do not indicate that the enzyme reactions
are irreversible
dicarboxylate/4 hydroxybutyrate cycle
Sato & Atomi, 2010
39
Kono et al., 2016
Reductive hexulose-phosphate pathway
Kono et al., 2016
Reductive hexulose-phosphate pathway
Structural comparison of archaeal and photosynthetic PRKs. (a,b) Topology diagrams of the folding patterns in protomers of (a) M. hungatei PRK (MhPRK) and (b) R. sphaeroides PRK (RsPRK) are shown in yellow and
blue, respectively. a-helices are denoted by cylinders, b-sheets by arrows and connecting loops by lines. Positions in the sequence that start and end each major secondary structural element are shown. (c,d) Ribbon diagrams of
(c) MhPRK (Protein Data Bank (PDB) ID 5B3F) monomer and (d) RsPRK (PDB ID 1A7J) monomer. Sulphate ions are bound to the active site of MhPRK and RsPRK. Disordered regions of missing electron density are shown as
dots for residues for 156–163 in MhPRK, residues 15–17 corresponding to a part of the P-loop, and residues 100–105 in RsPRK. (e,f) Active-site structures of (e) MhPRK and (f) RsPRK. Side chains of residues involved in ATPand
Ru5P-binding are shown as pink and green sticks, respectively, with oxygen atoms in red and nitrogen atoms in blue. Sulphate ions are shown in MhPRK and RsPRK, with sulphur atoms in green, and oxygen and nitrogen
atoms in the same colours as those in residue side chains. Small red balls represent water molecules. Disordered regions are shown as dots, and P-loop containing residues involved in ATP binding are shown in pink. Dotted lines
show interactions of active site sulphate ions. Kono et al., 2016
Reductive hexulose-phosphate pathway
Kono et al., 2016
Reductive hexulose-phosphate pathway
Kono et al., 2016
Reductive hexulose-phosphate pathway
Bar-Even et al., 2012
CO2 fixation pathways
45
Oxygen, metals and supply of C1 compounds
Autotrophic Euryarchaeota are strictly confined to anoxic conditions, generally specialized in
metabolizing C1 compounds and/or acetate, and their energy metabolism has low ATP yields.
Therefore, they need much of the C1-transforming machinery for their energy metabolism. The
reductive acetyl-coenzyme A (acetyl-CoA) pathway ideally copes with such constraints. Also,
essential metals are more available under anoxic conditions owing to the higher solubility of the
reduced forms of most metals.
In Crenarchaeota, the oxygen-sensitive dicarboxylate–hydroxybutyrate cycle is restricted to the
anaerobic Thermoproteales and Desulfurococcales, whereas the oxygen-insensitive
hydroxypropionate– hydroxybutyrate cycle is restricted to the mostly aerobic Sulfolobales and
Thaumarchaeota. The two lifestyles presuppose different electron donors with different redox
potentials and different oxygen sensitivity of cofactors and enzymes.
In a nutshell, energy cost-effective but oxygen-sensitive mechanisms cannot exist in aerobes
because the enzymes would be inactivated by oxygen; and not all anaerobes have C1
substrates at their disposal.
Berg et al., 2010
Benefits of the different autotrophic pathways
46
Energy demands
The different pathways require different amounts of ATP to make the cellular precursor
metabolites. The costs for synthesizing all auxiliary, CO2 fixation-related enzymes also differ,
which might determine the energy costs involved. The synthesis of the catalysts itself can
require a huge amount of energy as well as nitrogen and sulphur sources, especially if the
pathways involve many auxiliary enzymes. Carboxylases with low catalytic efficiency must be
synthesized in large amounts, as is the case for ribulose 1,5-bisphosphate carboxylase–
oxygenase (RUBISCO). So, energy limitation exerts a strong selective pressure in favour of
energy-saving mechanisms, and the energy costs are largely spent for the synthesis of
autotrophy-related enzymes. Berg et al., 2010
Benefits of the different autotrophic pathways
Könnicke et al., 2015
47
Properties of the CO2 fixation pathways
Berg et al., 2010; Bar-Even et al., 2012
48
Metabolic fluxes
In bacteria and archaea, the need for sugar phosphates in the biosynthesis of cell walls is lower
than in plants, which also synthesize huge amounts of cellulose and lignin that is derived from
erythrose 4-phosphate and phosphoenolpyruvate (PEP). The main metabolic fluxes are diverted
from acetyl-CoA, pyruvate, oxaloacetate and 2-oxoglutarate, and their synthesis from 3phosphoglycerate
is partly connected with a loss of CO2. Therefore, in bacteria autotrophic
pathways directly yielding acetyl-CoA are more economical. Still, most facultative aerobic
bacteria use the Calvin cycle, the regulation of which is almost detached from the central carbon
metabolism and therefore may be particularly robust.
Berg et al., 2010
Benefits of the different autotrophic pathways
49
CO2 species
As the bicarbonate (HCO3
–) concentration in slightly alkaline water is much higher than the
concentration of dissolved CO2, autotrophs might profit from using bicarbonate instead of CO2.
The usage of bicarbonate is a special feature of PEP carboxylase and biotin-dependent
carboxylases (that is, acetyl-CoA–propionyl-CoA carboxylase).
This property of PEP carboxylase is used in plants in the crassulacean acid and C4 metabolism
to increase the efficiency of photosynthesis. The same might be true for acetyl-CoA–propionylCoA
carboxylase, and the higher bicarbonate concentration could potentially make up for a
lower bicarbonate affinity.
Berg et al., 2010
Benefits of the different autotrophic pathways
50
Co-assimilation of organic compounds
Many autotrophic bacteria and archaea living in aquatic habitats probably encounter carbon
oligotrophic conditions and grow as mixotrophs. Co-assimilation of traces of organic compounds
might pay off. A complete or even a rudimentary hydroxypropionate–hydroxybutyrate cycle, for
instance, allows the co-assimilation of numerous compounds. These include fermentation
products and 3-hydroxypropionate, an intermediate in the metabolism of the ubiquitous
osmoprotectant dimethylsulphoniopropionate. It is possible that various widespread marine
aerobic phototrophic bacteria have genes encoding a rudimentary 3-hydroxypropionate cycle for
that purpose. Similarly, the dicarboxylate– hydroxybutyrate cycle allows the co-assimilation of
dicarboxylic acids and substrates that are metabolized through acetyl-CoA.
Berg et al., 2010
Benefits of the different autotrophic pathways
51
Microbial formate utilization
Crable et al. , 2011
52
Bar-Even, 2016
Microbial formate utilization
53
Microbial formate utilization
Reaction conditions: pH 7,0; 1M concentration, gases at 1 atm
Schink et al., 2017
54
Microbial formate utilization
Schink et al., 2017
55
Microbial formate utilization
Schink et al., 2017
56
Bar-Even et al., 2012
Microbial formate utilization
57
Microbial formate utilization
Bar-Even, 2016
Reductive acetyl-CoA pathway
58
Microbial formate utilization
Bar-Even, 2016
Serine cycle
59
Microbial formate utilization
Bar-Even, 2016
Reductive glycine pathway
60
61
Kottenhahn et al., 2018
H2 production from formate
Microbial formate utilization
62
Ceccaldi et al., 2017
A. woodii
63
Kottenhahn et al., 2018
H2 production from formate
64
Kottenhahn et al., 2018
H2 production from formate
Rittmann et al. , 2015
Microbial formate utilization
65
66
Rittmann et al. , 2015
Microbial formate utilization
67
Kim et al., 2010
H2 production from formate
68
Rittmann et al. (2012) Microbial Cell Factories
Rittmann et al. (2015) Biotechnology Advances
Ergal et al. (2018) Biotechnology Advances
HER: hydrogen evolution rate
H2 production from formate
Costa et al., 2010
Microbial formate utilization
69
Microbial formate utilization
Costa et al., 2013(c)
70
Microbial formate utilization
Costa et al., 2013(a,b)
71
Microbial formate utilization
Lupa et al., 2008
72
Microbial formate utilization
Lupa et al. , 2008
73
Microbial formate utilization
Crable et al. , 2011
74
C-cycle
Offre et al., 2014
Red arrows: metabolic stepts related to archaea and bacteria; Organge arrows: exclusive metabolic routes only related to arcahea
Modes of microbial anaerobic methane oxidation. There are four known ways in which microorganisms achieve anaerobic oxidation of methane (AOM). Two of these (a, b) are thought to rely on
obligate associations between two or more microbial partners, one of which performs oxidation and the other reduction; in the other two cases (c, d), a single microorganism performs both reactions.
a, Anaerobic methanotrophic archaea (ANMEs) oxidize methane (CH4) and convert it to carbon dioxide and water, in cooperation with sulphate-reducing bacteria, which convert sulphate (SO42−)
to hydrogen sulphide (H2S). The mechanism of energy exchange between the ANMEs and the sulphate-reducing bacteria is unknown. b, The oxidation of methane to CO2 by ANMEs is coupled
to the reduction of metal oxides, whereby metals such as manganese (Mn) or iron (Fe) are reduced to the +2 oxidation state. c, The bacterium Methoxymirabilis oxyfera converts nitrite (NO2−) to
nitric oxide (NO) and then dismutates (splits) NO into nitrogen and oxygen as diatomic gases. The bacterium then uses the resulting O2 to support methane oxidation. d, Milucka et al. show that some
ANMEs oxidize methane (as in a) but also reduce sulphate to zero-valent sulphur (S0), which they produce in the form of disulphide (HS2−). The disulphide can be used by associated bacteria,
Deltaproteobacteria, to yield sulphide (HS−) and sulphate, but this is an association of convenience, rather than necessity. Joye, 2012
Microbial methane oxidation
Microbial methane oxidation
Milucka et al., 2008
Disulphide disproportionation by Isis
enrichment culture.
a, Linear correlation between sulphate
and sulphide production in the Isis
enrichment culture incubated with 13CO2
and colloidal sulphur in the absence of
methane over the course of 70 days
(n=51) follows the 7:1 ratio inferred for
disulphide disproportionation.
b, Overlay of nanoSIMS12C14Nimage
(red) and fluorescence image of
microorganisms from the enrichment
culture stained with the DSS-658 probe
(green).
c, The 13C/12C image shows that the
DSS cells (several of which are shown by
white outlines) are enriched in 13C.
Coloured scale below panel c shows ratio
of the respective ions.
d, Production of sulphide and sulphate
(based on 35S accumulation in sulphate
after the addition of 35Ssulphide) under
AOM conditions (n=51; this experiment
was repeated twice).
Microbial methane oxidation
Milucka et al., 2008
Microbial methane oxidation
Revised model of anaerobic oxidation of
methane coupled to sulphate reduction.
ANME-2 oxidize methane with a concomitant
reduction of sulphate to zero-valent sulphur (S0,
elemental sulphur) that is partially deposited or
bound intracellularly. Produced S0 is exported or
diffuses outside the cell where it reacts with
sulphide to form polysulphides (disulphide,
among others).
Disulphide is taken up by the associated
Deltaproteobacteria and is disproportionated to
sulphate and sulphide. Sulphate produced during
disproportionation might be re-used by the
ANME and the ANME may also reduce some of
the sulphate all the way to sulphide (grey dotted
line). Dark circles in the bacteria represent
intracellular precipitates rich in iron and
phosphorus.
Milucka et al., 2008
Microbial methane oxidation
Significant pathways of
Methylomirabilis oxyfera.
Canonical pathways of denitrification (a),
aerobic methane oxidation (b) and
proposed pathway of methane oxidation
with nitrite (c).
narGHJI, nitrate reductase; napABCDE,
periplasmic nitrate reductase;
nirSJFD/GH/L, nitrite reductase; norZ,
nitric oxide reductase; nosDFYLZ, nitrous
oxide reductase; pmoCAB, particulate
methane monooxygenase;
mxaFJGIRSACKL/DE, methanol
dehydrogenase; fae, formaldehydeactivating
enzyme; mtdB,
methylenetetrahydromethanopterin
(H4MPT) dehydrogenase; mch, methenylH4MPT
cyclohydrolase; fhcABCD,
formyltransferase/hydrolase; fdhABC,
formate dehydrogenase.
Genes in red are absent from the
genome, those in blue are present in
the genome and those genes in green
are present in both the proteome and
the genome.
Asterisk, H4MPT-dependent reactions
involve the intermediates methylene
H4MPT, methenyl-H4MPT and formyl-
H4MPT.
Ettwig et al. , 2010
Microbial methane oxidation
Ettwig et al. , 2010
Thauer and Shima, 2006
Haroon et al. , 2013
Microbial methane oxidation
Haroon et al. , 2013
Key carbon and nitrogen
transformations in
‘Methanoperedens nitroreducens’.
Reverse methanogenesis pathway in
Ca. ‘M. nitroreducens’ coupled by an
unknown electron carrier to nitrate
reduction. Highly expressed genes are
shown in red, indicating that the
complete reverse methanogenesis
pathway and nitrate reduction genes
were active in the bioreactor.
Increasing line thickness indicates
increasing absolute gene expression
values. FPKG (fragments mapped per
kilobase of gene length) is a measure
of normalized gene expression.
Microbial methane oxidation
Haroon et al. , 2013
Microbial methane oxidation
Haroon et al. , 2013
Microbial methane oxidation