Metabolic modelling of syntrophic-like growth of a 1,3-propanediol producer, Clostridium butyricum, and a methanogenic archeon, Methanosarcina mazei, under anaerobic conditions

Abstract

Clostridium butyricum can convert glycerol into 1,3-propanediol, thereby generating unfortunately a high amount of acetate, formate and butyrate as inhibiting by-products. We have proposed a novel mixed culture comprising C. butyricum and a methane bacterium, Methanosarcina mazei, to relieve the inhibition and to utilise the by-products for energy production. In order to examine the efficiency of such a mixed culture, metabolic modelling of the culture system was performed in this work. The metabolic networks for the organisms were reconstructed from genomic and physiological data. Several scenarios were analysed to examine the preference of M. mazei in scavenging acetate and formate under conditions of different substrate availability, including methanol as a co-substrate, since it may exist in glycerol solution from biodiesel production. The calculations revealed that if methanol is present, the methane production can increase by 130%. M. mazei can scavenge over 70% of the acetate secreted by C. butyricum.

Appendices

Abbreviations used in the graphs, tables and text (for protein amino acids, standard abbreviations were used)

3-HPA:

3-Hydroxypropionaldehyde

3PG:

3-Phosphoglycerate

AcCoA, CH₃COCoA:

Acetyl-CoA

AKG:

α-Ketoglutarate

Asa:

l-Aspartate 4-semialdehyde

CH₂H₄MPT:

Methylenetetrahydromethanopterin

CH₃CoM:

Methyl coenzyme M

CH₃H₄MPT:

Methyltetrahydromethanopterin

CH₃H₄SPT:

Methyltetrahydrosarcinopterin

CHR:

Chorismate

CO:

Carbon monoxide

DHA:

Dihydroxyacetone

DHAP:

Dihydroxyacetonephosphate

E4P:

Erythrose-4-phosphate

EtOH:

Ethanol

F₄₂₀(H), F₄₂₀(red):

Coenzyme F₄₂₀ (reduced)

F6P:

Fructose-6-phosphate

FBP:

Fructose-1,6-biphosphate

Fd(H), FdH:

Ferredoxin (reduced)

FORM:

Formic acid (formate)

FUM:

Fumarate

G6P:

Glucose-6-phosphate

H₄MPT:

Tetrahydromethanopterin

H₄SPT:

Tetrahydrosarcinopterin

HAc:

Acetic acid (acetate)

HBu:

Butyric acid (butyrate)

HCH₄MPT:

Methenyltetrahydromethanopterin

HCOH₄MPT:

Formyltetrahydromethanopterin

Hse:

Homoserine

ICT:

Isocitrate

Ind:

Indole

kiV:

α-Ketoisovalerate

LAC:

Lactic acid (lactate)

MeOH:

Methanol

MethPhen(H), dHMePhe:

Methanophenazine (reduced)

OAA:

Oxalacetate

PEP:

Phosphoenolpyruvate

PPA:

Prephenate

PRPP:

Phosphoribosyl pyrophosphate

PYR:

Pyruvate

qCH₃COOH:

Specific acetate uptake rate

qCH₃OH:

Specific methanol uptake rate

qCH₄ :

Specific methane production rate

qCO₂ :

Specific carbon dioxide production/uptake rate

qH₂ :

Specific hydrogen uptake rate

qHCOOH:

Specific formate uptake rate

RIB5P:

Ribose-5-phosphate

S7P:

Sedoheptulose-7-phosphate

SKA:

Shikimate

SUC-CoA:

Succinyl-CoA

X5P:

Xylose-5-phosphate

${\text{Y}}_{{{\text{CH}}_{ 4} / {\text{CH}}_{ 3} {\text{COOH}}}}$ :

Methane to acetate yield coefficient

${\text{Y}}_{{{\text{CH}}_{ 4} / {\text{CH}}_{ 3} {\text{OH}}}}$ :

Methane to methanol yield coefficient

${\text{Y}}_{{{\text{CH}}_{ 4} / {\text{CO}}_{ 2} }}$ :

Methane to carbon dioxide yield coefficient

${\text{Y}}_{{{\text{CH}}_{ 4} / {\text{HCOOH}}}}$ :

Methane to formate yield coefficient

${\text{Y}}_{{{\text{CH}}_{ 4} / {\text{X}}}}$ :

Methane to biomass yield coefficient

${\text{Y}}_{{{\text{CO}}_{ 2} / {\text{H}}_{ 2} }}$ :

Carbon dioxide to hydrogen yield coefficient

${\text{Y}}_{{{\text{CO}}_{ 2} / {\text{HCOOH}}}}$ :

Carbon dioxide to formate yield coefficient

μ_MM :

M. mazei specific biomass growth rate

Abbreviations used in the formulation of metabolic network only (in the stoichiometric equations): exchanged metabolites

The notation was so designed that the names of all metabolites, which belong to C. butyricum start with letter “P” and so do the names of reaction rates listed down in Tables 1 and 2. For M. mazei, it is the letter “M”.

CO₂(exch):

Carbon dioxide (the exchanged pool)

FORM(exch):

Formate (the exchanged pool)

H₂(exch):

Hydrogen (the exchanged pool)

HAc(exch):

Acetate (the exchanged pool)

Intracellular metabolites of M. mazei

M_3PG:

3-Phosphoglycerate

M_AcCoA:

Acetyl coenzyme A

M_AcP:

Acetophosphate

M_aIVA:

α-Ketovalerate

M_AKG:

Aconitate

M_Ala:

Alanine

M_Arg:

Arginine

M_ASA:

l-Aspartate 4-semialdehyde

M_Asn:

Asparagine

M_Asp:

Aspartate

M_ATP:

Adenosinetriphosphate

M_CH₃CoM:

Methyl coenzyme M

M_CH₃H₄MPT:

Methyltetrahydromethanopterin

M_CH₃H₄SPT:

Methyltetrahydrosarcinopterin

M_CH₃OH:

Methanol

M_CH₄ :

Methane

M_CHR:

Chorismate

M_CO:

Carbon monoxide

M_CO₂ :

Carbon dioxide

M_Cys:

Cysteine

M_dHMethphen:

Methanophenazine (reduced)

M_DNA:

DNA

M_E4P:

Erythrose-4-phosphate

M_F₄₂₀red:

Coenzyme F₄₂₀ (reduced)

M_F6P:

Fructose-6-phosphate

M_FBP:

Fructose-1,6-diphosphate

M_FdH:

Ferredoxin (reduced)

M_FORM:

Formate

M_FORMH₄MPT:

Formyltetrahydromethanopterin

M_FUM:

Fumarate

M_G1P:

Glucose

M_G6P:

Glucose-6-phosphate

M_GA3P:

Glyceraldehyde phosphate

M_Gln:

Glutamine

M_Glu:

Glutamate

M_Gly:

Glycine

M_gly:

Glycogen

M_H⁺ :

Hydrogen ions

M_H₂ :

Hydrogen

M_HAc:

Acetic acid

M_His:

Histidine

M_Hse:

Homoserine

M_Ile:

Isoleucine

M_IND:

Indole

M_Leu:

Leucine

M_Lys:

Lysine

M_MeH₄MPT:

Methylenetetrahydromethanopterin

M_Met:

Methionine

M_MthH₄MPT:

Methenyltetrahydromethanopterin

M_N₂ :

Nitrogen

M_NADH:

NADH

M_NADPH:

NADPH

M_NH₄ ⁺ :

Ammonium ions

M_OAA:

Oxalacetate

M_PEP:

Phosphoenolpyruvate

M_Phe:

Phenylalanine

M_PLIP:

Phospholipids

M_PPA:

Prephenate

M_Pro:

Proline

M_PROT:

Protein

M_PRPP:

Phosphoribosyl pyrophosphate

M_PYR:

Pyruvate

M_RIB5P:

Ribose-5-phosphate

M_RNA:

RNA

M_SED7P:

Sedoheptulose-7-phosphate

M_Ser:

Serine

M_SKA:

Shikimate

M_SUCC_CoA:

Succinyl coenzyme A

M_Thr:

Threonine

M_Trp:

Tryptophane

M_Tyr:

Tyrosine

M_Val:

Valine

M_X:

Biomass

M_XYL5P:

Xylose-5-phosphate

Intracellular metabolites of C. butyricum

P_13PD:

1,3-Propanediol

P_3HPA:

3-Hydroxypropionealdehyde

P_AcCoA:

Acetyl coenzyme A

P_AKG:

α-ketoglutarate

P_ATP:

ATP

P_CO₂ :

Dioxide

P_DHA:

Dihydroxyacetone

P_DHAP:

Dihydroxyacetonephosphate

P_E4P:

Erythrose-4-phosphate

P_EtOH:

Ethanol

P_FdH:

Ferredoxin (reduced)

P_FORM:

Formate

P_FRU6P:

Fructose-6-phosphate

P_GA3P:

Glyceraldehyde-3-phosphate

P_GLC:

Glycerol

P_GLU6P:

Glucose-6-phosphate

P_H₂ :

Hydrogen

P_HAc:

Acetic acid

P_HBu:

Butyric acid

P_ISOCIT:

Isocitrate

P_LAC:

Lactate

P_NADH:

NADH

P_NADPH:

NADPH

P_NH₄ ⁺ :

Ammonium ions

P_OAA:

Oxalacetate

P_PEP:

Phosphoenolpyruvate

P_PYR:

Pyruvate

P_RIBOS5P:

Ribose-5-phosphate

P_RIBUL5P:

Ribulose-5-phosphate

P_S7P:

Sedoheptulose-7-phosphate

P_X:

Biomass

P_XYL5P:

Xylose-5-phosphate