Download msb4100131-sup-0001 - Molecular Systems Biology

Supplementary information
Development and characterization of syntrophic association
After stable syntrophic cultures were established in B3n media, we proceeded to characterize the
relationships between community growth and composition, and the metabolic interactions
underlying these relationships. The optical density, protein concentration, cell ratio, rRNA ratio,
and concentrations of metabolites in four cultures, grown in 240 ml bottles as described in
Material and methods, were measured from the time of inoculation and throughout exponential
growth and stationary phase. Results of this characterization are shown in Figures S1 and S2.
The syntrophic culture reproducibly converted 5.5 mmoles of lactate into 5 mmoles of acetate,
2.5 mmoles of methane and also carbon dioxide within about 123 hours of growth. Hydrogen
was produced within the first 50 hours of growth and then consumed to the minimal detectable
level by 176 hours. This hydrogen production pattern resembled the ‘burst’ which has been
observed in growth of D. vulgaris with sulfate (Noguera et al, 1998; Tsuji & Yagi, 1980).
Similar results were obtained in another experiment performed in tubes, and the same
stoichiometric relationships have been observed in chemostats (data not shown). The rate of
conversion of lactate into acetate, methane, and carbon dioxide, and the rate of change in
hydrogen concentration was calculated from these data and used to evaluate the flux-balance
model, as described in subsequent sections.
Population dynamic and 16S rRNA ratios of species in growing co-culture.
Because two different populations are involved in the production of acetate and methane from
lactate, we wanted to know the relationship between these metabolic processes and the relative
1
number and metabolic activity-level of the populations of each species. The relative proportion
of intact cells contributing to total community biomass was determined by microscopic counting
of fixed, fluorescently stained cells at several time intervals. As shown in Figure S1b, the
number of D. vulgaris cells was initially 2.5-fold higher than the number of M. maripaludis cells.
After a slight increase at 69 hours, the numerical dominance of D. vulgaris decreased gradually
throughout the experiment.
To assess the relative metabolic activity of each species, we compared the relative amount of 16S
rRNA contributed by the populations of each species over time. In general, cells tend to have
higher cellular rRNA when they are growing rapidly (Nomura, et al, 1984; Schaechter, et al,
1958). The amount of 16S rRNA produced by each species over time was measured using
capillary electrophoresis (Fig. S1c). Unlike the cell ratios, the ratio of 16S rRNA varied
significantly at different stages of growth in batch culture indicating that the growth rate of each
species varied throughout batch culture growth. The inoculum used to start the growth
experiment came from a culture that had reached stationary phase more than one day earlier.
Therefore, the ratio of 16S rRNA at time zero likely reflects a stationary-phase physiological
state. At this time point, the concentration of M. maripaludis 16S rRNA was approximately 2fold greater than that of D. vulgaris, despite the fact that the number of M. maripaludis cells was
about 2.5-fold less. These results indicate that the rRNA content per cell must have been
between 3 and 5-fold higher in M. maripaludis than in D. vulgaris depending on the percentage
of intact D. vulgaris cells that are viable and contain rRNA (70-100% viable, respectively).
Between 0 and 50 hr of growth, however, the relative population activity level of the two species
reversed, with D. vulgaris producing about 1.7-fold more 16S rRNA overall than M.
2
maripaludis, and this relationship remained constant until after 100 hours of growth. This ratio
is still lower than the ratio of cell types, indicating that M. maripaludis still maintains more
rRNA per cell than D. vulgaris, although the difference must be smaller than at time zero. After
100 hours of growth, the ratio of 16S rRNA decreased again until the ratio of rRNA production
was only slightly less than one (0.9), indicating that either the activity level of M. maripaludis
increased, the activity level of D. vulgaris decreased, or both.
Analysis of metabolites in growth medium.
In addition to the growth and community dynamics studies presented above, we also explored
the possibility that other metabolites could be secreted by D. vulgaris during syntrophic growth.
Analysis of the D. vulgaris genome sequence together with experimental data suggested that
ethanol, glycerol, formate, succinate, and fumarate may be produced in the culture medium. We
tested for the presence of these compounds in the medium using chromatographic and enzymatic
methods. Fumarate, formate, glycerol and succinate were not detected under the typical growth
conditions, suggesting that if they were present, their concentration would have to be below
0.1mM (lower limit of detection). Approximately 20 - 100 umole of ethanol was detected in
batch and chemostat co-cultures grown at steady state.
Growth stoichiometry of M maripaludis monoculture.
To assess whether the M. maripaludis model is capable of accurately predicting the relationship
between growth rate and uptake of substrates, we measured hydrogen, acetate, and carbon
dioxide uptake, methane and protein production over time for three independent cultures of M.
maripaludis growing on hydrogen, acetate, and carbon dioxide. Figure S3 shows the change in
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hydrogen, acetate, methane, and carbon dioxide over time. In our experiment 160 (± 20 based on
standard deviation, n=3) micromoles of acetate was consumed along with hydrogen and carbon
dioxide.
Appendix A: List of Desulfovibrio vulgaris reactions included in the model. Associated
enzymes with putative genes found in the genome annotation are listed below each reaction.
Cases where one or more genes are not found are also noted. Note that some reactions are
lumped, and thus represent multiple enzymatic steps.
1.
SO4(-2) + ATP + H+(ext) APS + PPi
Sulfate adenylyltransferase
2.
APS + Cyt-C(red)  SO3(-2) + Cyt-C(ox) + AMP
Adenylylsulfate kinase; 3’-phosphoadenylylsulfate reductase
3.
H2 + Cyt-C(ox)  Cyt-C(red) + 2 H+(ext)
Cytochrome C3-reducing hydrogenase
4.
H2 + Fdx(ox)  Fdx(red)
Ferredoxin-reducing hydrogenase
5.
SO3(-2) + 3 Cyt-C(red)  H2S + 3 Cyt-C(ox) + 6 H+(ext)
Sulfite reductases (varying electron donors)
6.
NADH + Q  NAD + QH2
NADH dehydrogenase (non-proton translocating)
7.
Fdx(red) + Q  Fdx(ox) + QH2
8.
QH2 + Cyt-C(ox)  Cyt-C(red) + Q
9.
2 H+(ext) + ADP  ATP
ATP synthase
10.
ATP + AMP  2 ADP
adenylate kinase
11.
Lac + Fdx(ox)  Pyr + Fdx(red)
L/D lactate dehydrogenase
12.
Lac + NAD  Pyr + NADH
4
L/D lactate dehydrogenase
13.
Pyr + Fdx(ox)  AcCoA + CO2 + Fdx(red)
pyruvate ferrodoxin oxidoreductase
14.
G6P  F6P
Glucose-6P isomerase
15.
G6P + NADP  6PGL + NADPH
Not found: glucose-6P dehydrogenase
16.
F6P + ATP  FBP +ADP
PPi-Fructose-6P 1-phosphotransferase (PFK)
17.
FBP  F6P + Pi
Fructose-1,6-bisphosphatase
18.
FBP  2 GAP
Fructose bisphosphate aldolase
19.
X5P + R5P  Sed7P + GAP
transketolase, transaldolase (putative)
20.
Sed7P + GAP  E4P + F6P
transketolase, transaldolase (putative)
21.
X5P + E4P  F6P + GAP
transketolase, transaldolase (putative)
22.
Ribu5P  R5P
ribose 5-phosphate isomerase B
23.
Ribu5P X5P
ribulose-phosphate 3-epimerase
24.
GAP + NAD +ADP  3PG + NADH + ATP
Glyceraldehyde-3P dehydrogenase; phosphoglycerate kinase
25.
3PG  PEP
phosphoglycerate mutase; enolase
26.
PEP + ADP  Pyr + ATP
pyruvate kinase
27.
Pyr + ATP  PEP + AMP
Phosphoenolpyruvate synthase
5
28.
Pyr + ATP + CO2  OAA + ADP
pyruvate carboxylase
29.
AcCoA  AcP + CoA
phosphate acetyl transferase
30.
AcP + ADP  AcOH + ATP
acetate kinase
31.
AcCoA + OAA  Cit + CoA
not found: citrate synthase
32.
AcOH + ATP + CoA  ATP + AcCoA
acetyl-CoA synthase
33.
Cit  Isocit
aconitase
34.
Isocit + NAD  KG + NADPH + CO2
isocitrate dehydrogenase (NADP-linked)
35.
SucCoA + ADP  Suc + ATP + CoA
succinyl-CoA synthetase
36.
Suc + FAD  Fum + FADH
succinate dehydrogenase/fumarate reductase (3 subunits)
37.
Fum  Mal
fumarate hydratase (classes I and II present)
38.
Mal + NADP  Pyr + CO2 + NADPH
malic enzyme (NADP-linked)
39.
KG + NADH + NH3  Glu + NAD
glutamate dehydrogenase (NAD-linked)
40.
KG + NADPH + Gln  2 Glu + NADP
glutamate synthase (NADP-linked)
41.
Glu + ATP + NH3  Gln + ADP
glutamine synthetase
42.
OAA + Glu  Asp + KG
aspartate aminotransferase
43.
Asp + 2 ATP + NH3  Asn + 2 ADP
6
aspartate-ammonia lyase
44.
Asp + ATP + NADPH  ASA + ADP + NADP
aspartate kinase, aspartate semialdehyde dehydrogenase
45.
ASA + Pyr  DDP
dihydrodipicolinate synthase
46.
DDP + NADPH  THDP + NADP
dihydrodipicolinate reductase
47.
THDP + SucCoA + Glu  SDAP + KG + CoA
not found: dapC, dapD
48.
SDAP  mDAP + Suc
succinyl-diaminopimelate desuccinylase; diaminopimelate epimerase
49.
mDAP  Lys + CO2
diaminopimelate decarboxylase
50.
Glu + 2NADPH + ATP  Pro + 2NADP + ADP
glutamate-5-kinase
gamma-glutamyl phosphate reductase
pyrroline-5-carboxylate reductase
51.
Glu + 5ATP + NADPH + Gln + Asp + AcCoA + CO2  Arg + 5ADP + NADP + KG +
Fum
glutamate N-acetyltransferase/amino-acid acetyltransferase
argininosuccinate lyase
argininosuccinate synthase
ornithine carbamoyltransferase
acetylglutamate kinase
possible acetylornithine transaminase
N-acetyl-gamma-glutamyl-phosphate reductase
carbamoyl-phosphate synthase, both subunits
Not found: acetylornithine deacetylase
52.
ASA + NADPH  Hse + NADP
homoserine dehydrogenase
53.
Thr + Glu + NADPH + Pyr  Ile + KG + NADP + NH3 + CO2
7
acetolactate synthase, both subunits
ketol-acid reductoisomerase
branched-chain amino acid aminotransferase
dihydroxy-acid dehydratase
Not found: threonine dehydratase
54.
Ser + H4F  Gly + methylene-H4F
Serine hydroxymethyltransferase
55.
methylene-H4F + NADP  NADPH + H4F + Form
formyl-H4F deformylase, methylene-H4F dehydrogenase
methenyl-H4F cyclohydrolase
56.
NAD + Gly + H4F  methylene-H4F + NADH + CO2 + NH3
glycine cleavage system (lpdA, gcvPB, gcvPA, gcvH)
57.
PEP + E4P + NADPH  SKA + NADP
3-dehydroquinate dehydratase, type II; shikimate 5-dehydrogenase
Not found: dehydroquinate synthase, DHAP synthase
58.
SKA + PEP + ATP  CHR + ADP
chorismate synthase
3-phosphoshikimate 1-carboxyvinyltransferase
shikimate kinase
59.
CHR  PPA
chorismate mutase (putative)
60.
PPA + NAD + Glu  Tyr + NADH + CO2 + KG
prephenate dehydrogenase; aromatic amino acid transaminase
61.
PPA + Glu  Phe + CO2 + KG
prephenate dehydrogenase; aromatic amino acid transaminase
62.
CHR + R5P + 2 ATP + Gln  Ind + Glu + Pyr + CO2 + GAP +2ADP
anthranilate phosphoribosyltransferase
anthranilate synthase component I
indole-3-glycerol phosphate synthase
63.
Ind + Ser  Trp
tryptophan synthase, alpha and beta subunits
8
64.
2 Pyr  ALC
acetolactate synthase, both subunits
65.
ALC + NADPH  IVA + NADP + CO2
ketol-acid reductoisomerase; dihydroxy-acid dehydratase
66.
IVA + Glu  Val + KG
branched-chain amino acid aminotransferase
67.
Val + Pyr  Ala + aIVA
alanine racemase, transaminase (putative)
68.
aIVA + AcCoA + NAD + Glu  Leu + NADH + CO2 + KG + CoA
2-isopropylmalate synthase
3-isopropylmalate dehydrogenase
branched-chain amino acid aminotransferase
3-isopropylmalate dehydratase (isomerase)
69.
R5P + ATP  PRPP + AMP
70.
PRPP + 2 ATP + Gln + 2 NAD  His + 2 ADP + KG + 2 NADH
ribose-phosphate pyrophosphokinase
phosphoribosylformimino-5-aminoimidazole carboxamide ribotide isomerase
phosphoribosyl-AMP cyclohydrolase
ATP phosphoribosyltransferase
imidazoleglycerol-phosphate dehydratase
imidazoleglycerol phosphate synthase, both subunits
histidinol-phosphate aminotransferase
histidinol dehydrogenase
71.
Ser + AcCoA + H2S  Cys + AcOH
serine acetyltransferase; cysteine synthase A
72.
3PG  Ser
D-3-phosphoglycerate dehydrogenase
Not found: phosphoserine transaminase; phosphoserine phosphatase
73.
Hse + ATP  Thr + ADP
threonine synthase
Not found: Homoserine kinase
74.
Hse + SucCoA + Cys + methylene-H4F  Met + Suc + CoA + H4F + Pyr + NH3
9
5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase
5-methyltetrahydrofolate-homocysteine methyltransferase
Not found: 3 enzymes for converting homoserine to homocysteine
75.
AcCoA + Glyox  Mal + CoA
malate synthase
76.
Pyr + CoA  AcCoA + Form
pyruvate-formate lyase
77.
AcCoA + 2 NADH  Ethanol
aldehyde-alcohol dehydrogenase
78.
Form + Cyt-C(ox)  Cyt-C(red) + CO2
formate dehydrogenase
79.
Form  Form(ext)
80.
H2  H2(ext)
81.
GAP + FADH  glycerol-P + FAD
anaerobic glycerol-3P dehydrogenase
82.
glycerol-P  glycerol + ATP
glycerol kinase
83.
NH3  NH3(ext)
84.
AcOH  AcOH(ext)
85.
Lac(ext) + 0.5 ATP  Lac + 0.5 ADP
86.
KG + CoA + Fdx(ox)  SucCoA + Fdx(red)
2-oxoglutarate synthase
87.
6PGL  6PG
phosphoglucolactonase
88.
6PG + NADP  Ribu5P + NADPH
phosphoglucanate dehydrogenase
89.
Biomass Production:
RNA (21.33%)
10
3.47 PRPP + 5.02 Gln + -5.02 Glu + 3.08 Gly + 6.17 Asp + 32.41 ATP + -32.41 ADP + 6.17
methylene-H4F + -6.17 H4F + 3.09 NAD + -3.09 NADH + 6.17 NADP + -6.17 NADPH + 1.16
CO2 + -3.47 Fum + -3.86 NH3
DNA (3.23%)
3.37 PRPP + 4.88 Gln + -4.88 Glu + 3 Gly + 6 Asp + 31.5 ATP + -31.5 ADP + 7.12 mTHF + 7.12 THF + 3 NAD + -3 NADH + 3.75 NADP + -3.75 NADPH + 1.12 CO2 + -3.37 Fum + -3.75
NH3
Phospholipid (9.47%)
20.8 AcCoA + -20.8 CoA + 1.95 GAP + 0.65 Ser + 44.2 ATP + -44.2 ADP + 38.35 NADH + 38.35 NAD + -0.65 CO2
Peptidoglycan (2.60%)
1.94 F6P + 1.94 AcCoA + -1.94 CoA + 1.94 Gln + -1.94 Glu + 2.91 Ala + 0.97 PEP + 0.97 Lys
+ 6.97 ATP + -6.97 ADP + 0.97 NADPH + -0.97 NADP + -0.97 CO2
LPS (3.54%)
0.91 R5P + 0.91 F6P + 0.91 PEP + 15.47 AcCoA + -0.91 AcOH + -0.91 Glu + 0.91 Gln + 32.76
ATP + 12.74 NADH
Protein (57.23%)
0.77 Gly + 0.96 Ala + 0.67 Val + 0.85 Leu + 0.44 Ile + 0.44 Ser + 0.48 Thr + 0.30 Phe + 0.26
Tyr + 0.01 Trp + 0.15 Cys + 0.22 Met + 0.54 Lys + 0.46 Arg + 0.16 His + 0.46 Asp + 0.52 Glu +
0.46 Asn + 0.52 Gln + 0.34 Pro + 18 ATP + -18 ADP
Glycogen (2.60%)
F6P + ATP + -1 ADP
11
Appendix B: List of Methanococcus maripaludis reactions included in the model.
Associated enzymes with putative genes found in the genome annotation are listed below
each reaction. Cases where one or more genes are not found are also noted. Note that
some reactions are lumped, and thus represent multiple enzymatic steps.
1.
CO2 + Fdx(red) + MFR  Formyl-MFR + Fdx(ox)
Formyl-methanofuran dehydrogenase
2.
CO2 + CODH + Fdx(red)  CO-CODH + Fdx(ox)
Carbon monoxide dehydrogenase
3.
Formyl-MFR + H4MPT  MFR + Formyl-H4MPT
Formyl-tetrahydromethanopterin formyltransferase
4.
Formyl-H4MPT  Methenyl-H4MPT
Methenyl-tetrahydromethanopterin cyclohydrolase
5.
Methenyl-H4MPT + F420(red)  Methylene-H4MPT + F420(ox)
Methylene-tetrahydromethanopterin dehydrogenase (Coenzyme F420-dependent)
6.
Methenyl-H4MPT + H2  Methylene-H4MPT
Methylene-tetrahydromethanopterin dehydrogenase (H2-dependent)
7.
Methylene-H4MPT + F420(red)  Methyl-H4MPT + F420(ox)
Methylene-tetrahydromethanopterin reductase
8.
Methyl-H4MPT + B12  Methyl-B12
Methyltetrahydromethanopterin –coenzyme B12 methyltransferase
9.
Methyl-B12 + CO-CODH  Acetyl-CODH + B12
Carbon monoxide dehydrogenase
10.
Acetyl-CODH + CoA  AcCoA + CODH
Acetyl-CoA synthase
11.
Methyl-H4MPT + CoM-SH  CoM-SCH3 + 2 H+(ext)
Methyltetrahydromethanopterin –coenzyme M methyltransferase
12.
CoM-SCH3 + HS-HTP  CH4 + CoM-SSHTP
13.
H2 + CoM-SSHTP  HS-HTP + CoM-SH + 2 H+(ext)
Methyl coenzyme M reductase
14.
H2 + Fdx(ox) + 2 H+(ext)  Fdx(red)
12
ferredoxin-reducing hydrogenase
15.
H2(ext)  H2
16.
F420(ox) + H2  F420(red)
hydrogen uptake
F420-reducing hydrogenase
17.
NAD + F420(red)  NADH + F420(ox)
ferredoxin oxidoreductase
18.
NADP + F420(red)  NADPH + F420(ox)
NADP oxidoreductase
19.
Form + F420(ox)  F420(red) + CO2
formate dehydrogenase
20.
Form(ext)  Form
21.
3 H+(ext) + ADP  ATP
formate uptake
ATPase
22.
ATP + AMP  2 ADP
adenylate kinase
23.
Pyr + Fdx(ox)  AcCoA + CO2 + Fdx(red)
pyruvate ferrodoxin oxidoreductase
24. G6P  F6P
Glucose-6P isomerase
25. G6P  G1P
phosphoglucomutase
26. F6P + ADP  FBP +AMP
ADP-dependent phosphofructokinase
27. FBP  F6P + Pi
Fructose-1,6-bisphosphatase
28. FBP  2GAP
Fructose bisphosphate aldolase (putative)
29. X5P + R5P  Sed7P + GAP
30. Sed7P + GAP  E4P + F6P
31. X5P + E4P  F6P + GAP
Transaldolase
13
Not found: transketolase
32. GAP + NAD +ADP  3PG + NADH + ATP
Glyceraldehyde-3P dehydrogenase; phosphoglycerate kinase
33. GAP + Fdx(ox) + ADP  3PG + Fdx(red) + ATP
Glyceraldehyde-3P ferredoxin oxidoreductase
34. 3PG  PEP
phosphoglycerate mutase; enolase
35. PEP + ADP  Pyr + ATP
pyruvate kinase
36. Pyr + ATP  PEP + AMP
phosphoenolpyruvate synthase
37. Pyr + ATP + CO2  OAA + ADP
pyruvate carboxylase
38. AcOH + ATP + CoA  ADP + Acetyl-CoA
acetyl-CoA synthetase (AMP-forming and ADP-forming present)
39. SucCoA + Fdx(red) + CO2  -KG + Fdx(ox) + CoA
2-oxoglutarate ferredoxin oxidoreductase
40. SucCoA + ADP  Suc + ATP + CoA
succinate--CoA ligase (ADP-forming)- alpha and beta subunits
41. Suc + FAD  Fum + FADH
Succinate dehydrogenase/fumarate reductase (2 subunits)
42. Fum  Mal
fumarate hydratase
43. Mal + NAD  OAA +NADH
malate dehydrogenase (NADP)
44. KG + NADPH + Gln  2 Glu
glutamate synthase
45. Glu + ATP + NH3  Gln + ADP
glutamine synthetase
46. OAA + Glu  Asp + KG
aspartate aminotransferase
14
47. Asp + ATP + NH3  Asn + AMP
asparagine synthase
48. Asp + ATP + NADPH  ASA + ADP + NADP
aspartate kinase; aspartate semialdehyde dehydrogenase
49. ASA + Pyr  DDP
dihydrodipicolinate synthase
50. DDP + NADPH  THDP + NADP
dihydrodipicolinate reductase
51. THDP + SucCoA + Glu  SDAP + KG + CoA
Not found: dapC, dapD
52. SDAP  mDAP + Suc
succinyl-diaminopimelate desuccinylase; diaminopimelate epimerase
53. mDAP  Lys + CO2
diaminopimelate decarboxylase
54. Glu + 5ATP + NADPH + Gln + Asp + AcCoA + CO2  Arg + 5ADP + NADP +
KG + Fum
glutamate N-acetyltransferase/amino-acid acetyltransferase
argininosuccinate lyase
argininosuccinate synthase
ornithine carbamoyltransferase
acetylglutamate kinase
acetylornithine transaminase (possible)
N-acetyl-gamma-glutamyl-phosphate reductase
carbamoyl-phosphate synthase (both subunits found)
Not found: acetylornithine deacetylase
55. ASA + NADPH  Hse + NADP
homoserine dehydrogenase
56. Hse + ATP  Thr + ADP
threonine synthase; homoserine kinase
57. Pyr + AcCoA + CO2 + NH3  Ile
Uncharacterized pathway
15
58. 3PG  Ser
D-3-phosphoglycerate dehydrogenase
phosphoserine phosphatase
Not found: phosphoserine transaminase
59. PEP + 3PG + CO2  SKA
dehydroquinate synthase, DHAP synthase not found
3-dehydroquinate dehydratase, type I
shikimate 5-dehydrogenase
Not found: dehydroquinate synthase; DHAP synthase
60. SKA + PEP + ATP  CHR + ADP
chorismate synthase
3-phosphoshikimate 1-carboxyvinyltransferase
shikimate kinase
61. CHR  PPA
Not found: chorismate mutase
62. PPA + NAD + Glu  Tyr + NADH + CO2 + KG
prephenate dehydrogenase; aromatic amino acid transaminase (possible)
63. PPA + Glu  Phe + CO2 + aKG
prephenate dehydrogenase; aromatic amino acid transaminase (possible)
64. CHR + PRPP  Ind + Pyr + CO2 + GAP
anthranilate phosphoribosyltransferase
anthranilate synthase component I
indole-3-glycerol phosphate synthase
65. Ind + Ser  Trp
tryptophan synthase (alpha and beta subunits)
66. 2 Pyr  ALC
acetolactate synthase (both subunits)
67. ALC + NADPH  IVA + NADP + CO2
ketol-acid reductoisomerase; dihydroxy-acid dehydratase
68. IVA + Glu  Val + KG
branched-chain amino acid aminotransferase
16
69. Val + Pyr  Ala + IVA
alanine racemase
Not found: transaminase
70. IVA + AcCoA + NAD + Glu  Leu + NADH + CO2 + KG + CoA
2-isopropylmalate syntase; 3-isopropylmalate dehydratase
3-isopropylmalate dehydrogenase
branched-chain amino acid aminotransferase
71. R5P + ATP  PRPP + 2 AMP
ribose-phosphate pyrophosphokinase
72. PRPP + 2 ATP + Gln + 2 NAD  His + 2 ADP + aKG + 2 NADH
phosphoribosylformimino-5-aminoimidazole carboxamide ribotide isomerase
phosphoribosyl-AMP cyclohydrolase
ATP phosphoribosyltransferase
imidazoleglycerol-phosphate dehydratase
imidazoleglycerol phosphate synthase (both subunits)
histidinol-phosphate aminotransferase
histidinol dehydrogenase
73. Ser + H4MPT  Gly + mH4MPT
Not found: Serine hydroxymethyltransferase
74. Ser + AcCoA + H2S  Cys + AcOH
Not found: serine acetyltransferase; cysteine synthase
75. Glu + 2 NADPH + ATP  Pro + 2 NADP + ADP
glutamate-5-kinase (possible)
Not found: gamma-glutamyl phosphate reductase; pyrroline-5-carboxylate
reductase
76. Hse + SucCoA + Cys + methylene-H4F  Met + Suc + CoA + H4F + Pyr + NH3
5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase
homoserine dehydrogenase
Not found: 3 enzymes for converting homoserine to homocysteine
77. X5P  R5P
Not found: ribose 5-phosphate isomerase; ribulose-phosphate 3-epimerase
17
78. NH3(ext)  NH3
79. Ala(ext)  Ala
80. Ala + NAD  NADH + NH3 + Pyr
81. N2 + 4 Fdx(red) + 16 ATP  2 NH3 + 4 Fdx(ox) + 16 ADP + H2
Nitrogenase
82.
Biomass Production:
RNA (22.73%)
3.47 PRPP + 5.02 Gln + -5.02 Glu + 3.08 Gly + 6.17 Asp + 32.41 ATP + -32.41 ADP +
6.17 mTHF + -6.17 THF + 3.09 NAD + -3.09 NADH + 6.17 NADP + -6.17 NADPH +
1.16 CO2 + -3.47 Fum + -3.86 NH3
DNA (3.44%)
3.37 PRPP + 4.88 Gln + -4.88 Glu + 3 Gly + 6 Asp + 31.5 ATP + -31.5 ADP + 7.12
mTHF + -7.12 THF + 3 NAD + -3 NADH + 3.75 NADP + -3.75 NADPH + 1.12 CO2 + 3.37 Fum + -3.75 NH3
Phospholipid (10.09%)
20.8 AcCoA + -20.8 CoA + 1.95 GAP + 0.65 Ser + 44.2 ATP + -44.2 ADP + 38.35
NADH + -38.35 NAD + -0.65 CO2
Protein (60.97%)
0.77 Gly + 0.96 Ala + 0.67 Val + 0.85 Leu + 0.44 Ile + 0.44 Ser + 0.48 Thr + 0.30 Phe +
0.26 Tyr + 0.01 Trp + 0.15 Cys + 0.22 Met + 0.54 Lys + 0.46 Arg + 0.16 His + 0.46 Asp
+ 0.52 Glu + 0.46 Asn + 0.52 Gln + 0.34 Pro + 18 ATP + -18 ADP
Glycogen (2.77%)
F6P + ATP + -1 ADP
18
Appendix C: Abbreviation of metabolites
3PG
6PG
6PGL
AcCoA
AcOH
AcP
IVA
KG
ALC
APS
ASA
B12
CHR
Cit
CoA
CODH
CoM-SCH3
CoM-SH
CoM-SSHTP
Cyt-C(ox)
Cyt-C(red)
DDP
E4P
F420(ox)
F420(red)
F6P
FBP
Fdx(ox)
Fdx(red)
Form
FormylH4MPT
Formyl-MFR
Fum
G1P
G6P
GAP
Glycerol-P
Glyox
: 3-phospho-D-glycerate
: 6-phospho-D-gluconate
: 6-Phospho-D-glucono-1,5-lactone
: acetylcoenzyme-A
: acetate
: acetylphosphate
: alpha-keto-isovaleric acid
: 2-oxoglutaric acid
: acetohydroxy acid
: adenylylsulfate
: aspartate semialdehyde
: coenzyme B12
: chorismic acid
: citric acid
: coenzyme A
: carbon monoxide dehydrogenase
: methyl-coenzyme M
: coenzyme M
: cytochrome C3 (oxidized form)
: cytochrome C3 (reduced form)
: dihydrodipicolinate
: erythrose-4-phosphate
: coenzyme F420 (oxidized)
: coenzyme F420 (reduced)
: fructose-6-phosphate
: fructose bisphosphate
: ferredoxin (oxidized form)
: ferredoxin (reduced form)
: formate
: formyl-tetrahydromethanopterin
: flormy-methanofuran
: fumerate
: glucose-1-phosphate
: glucose-6-phosphate
: glyceraldehyde-phosphate
: glycerol phosphate
: glyoxylate
19
H+(ext)
H4F
H4MPT
HS-HTP
Hse
Ind
Isocit
Lac
Mal
mDAP
methenylH4MPT
methyl-B12
methyl-H4MPT
methyleneH4MPT
methylene-H4F
MFR
OAA
PEP
PPA
PRPP
Pyr
Q
QH2
R5P
Ribu5P
SDAP
SKA
Sed7P
SO4(-2)
SO3(-2)
Suc
SucCoA
THDP
X5P
: external proton
: tetrahydrofolate
: tetrahydromethanopterin
: homoserine
: indole glycerol phosphate
: isocitrate
: D/L-lactic acid
: malic acid
: meso-diaminopimelate
: methenyl-tetrahydromethanopterin
: methyl-coenzyme B12
: methyl-tetrahydromethanopterin
: methylene-tetrahydromethanopterin
: methylene tetrahydrofolate
: methanofuran
: oxaloacetate
: phosphoenolpyruvate
: prephenate
: phosphoribosyl pyrophosphate
: pyruvic acid
: ubiquinone (oxidized form)
: ubiquinone (reduced form)
: ribose-5-phosphate
: ribulose-5-phosphate
: n-succinyl-L-2,6-diaminoheptanedioate
: shikimate
: D-sedoheptulose-7-phosphate
: inorganic sulfate
: inorganic sulfite
: succinic acid
: succinyl-coenzyme A
: tetrahydrodipicolinate
: xylulose-5-phosphate
Commonly accepted abbreviations such as amino acids, common energy or redox
metabolites, and molecular formulas are not listed.
20