Literature summary for 1.2.7.1 extracted from

Lin, W.C.; Yang, Y.L.; Whitman, W.B.
The anabolic pyruvate oxidoreductase from Methanococcus maripaludis (2003), Arch. Microbiol., 179, 444-456.

General Stability

General Stability	Organism
following dialysis, the enzyme is very unstable in the absence of glycerol or ethylene glycol, thiamine diphosphate and MgCl2 also help to maintain activity	Methanococcus maripaludis

Inhibitors

Inhibitors	Comment	Organism
Dithionite	with 5 mM dithionite, the half-life is 7 h at 2 °C, with about 90% of the original activity being lost after 24 h	Methanococcus maripaludis
glyoxylate	20 mM, about 40% of the original activity is lost. 2 mM glyoxylate inhibits 6%	Methanococcus maripaludis
additional information	no substrate inhibition with CoA up to 0.1 mM. Slightly or not inhibited at all by glyoxylate, nitrite, CO or potential physiological effectors. Not inhibited by CO. Potential affectors such as ATP, ADP, AMP, cAMP, GTP, GDP, GMP, NAD+, NADH, and glyceraldehyde 3-phosphate, at concentrations of 2 mM, do not affect the activity	Methanococcus maripaludis
oxygen	-	Methanococcus maripaludis

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
0.0058	-	CoA	pH 8.6, 37°C	Methanococcus maripaludis
0.115	-	pyruvate	pH 8.6, 37°C	Methanococcus maripaludis
0.205	-	2-oxobutyrate	pH 8.6, 37°C	Methanococcus maripaludis
0.264	-	oxaloacetate	pH 8.6, 37°C	Methanococcus maripaludis

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Iron-sulfur cluster	porE encodes the 21500 Da subunit that contains a high cysteinyl residue content and a motif indicative of a [FeS] cluster. Subunit porF also also has a high cysteinyl residue content, and two [FeS] cluster motifs. Based upon these results, it is proposed that PorE and PorF are components of a specialized system required to transfer low-potential electrons for pyruvate biosynthesis	Methanococcus maripaludis

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
190000	-	-	Methanococcus maripaludis

Organism

Organism	UniProt	Comment	Textmining
Methanococcus maripaludis	Q9P9E5 and Q9P9E4 and Q9P9E6 and Q9P9E7 and Q9P9E3	Q9P9E5: subunit alpha, Q9P9E4: subunit beta, Q9P9E6: subunit gamma, Q9P9E7: subunit delta, Q9P9E3: subunit PorE	-
Methanococcus maripaludis DSM Z 2067	Q9P9E5 and Q9P9E4 and Q9P9E6 and Q9P9E7 and Q9P9E3	Q9P9E5: subunit alpha, Q9P9E4: subunit beta, Q9P9E6: subunit gamma, Q9P9E7: subunit delta, Q9P9E3: subunit PorE	-

Oxidation Stability

Oxidation Stability	Organism
the enzyme is very sensitive to O2. Following incubation in air at 2°C for 40 min, about 60% of enzyme activity is lost. The half-life is 5.2 min when the purified enzyme is exposed to air in an ice bath. After inactivation of the purified enzyme by oxygen, activity is not restored byreplacing the oxygen with nitrogen and adding 0.01 mM dithionite no activity is lost after dialysis of extract in a basic buffer containing 20 mM potassium Tricine, pH 8.6, 5 mM MgCl2, 0.5 mM dithiothreitol, 0.1 mM thiamine diphosphate, and 10% glycerol	Methanococcus maripaludis

Organism

the enzyme is very sensitive to O2. Following incubation in air at 2°C for 40 min, about 60% of enzyme activity is lost. The half-life is 5.2 min when the purified enzyme is exposed to air in an ice bath. After inactivation of the purified enzyme by oxygen, activity is not restored byreplacing the oxygen with nitrogen and adding 0.01 mM dithionite no activity is lost after dialysis of extract in a basic buffer containing 20 mM potassium Tricine, pH 8.6, 5 mM MgCl2, 0.5 mM dithiothreitol, 0.1 mM thiamine diphosphate, and 10% glycerol

Methanococcus maripaludis

Purification (Commentary)

Purification (Comment)	Organism
-	Methanococcus maripaludis

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
0.3	-	pH 8.6, 37°C, substrate: indol-3-pyruvate	Methanococcus maripaludis
3.6	-	pH 8.6, 37°C, substrate: 2-oxobutyrate	Methanococcus maripaludis
6.5	-	pH 8.6, 37°C, substrate: oxaloacetate	Methanococcus maripaludis
7.4	-	pH 8.6, 37°C, substrate: pyruvate	Methanococcus maripaludis

Storage Stability

Storage Stability	Organism
-20°C, 3 weeks, anaerobic conditions, no loss of activity of the partially purified enzyme	Methanococcus maripaludis
2°C, 2 weeks, anaerobic conditions, no loss of activity	Methanococcus maripaludis

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
2-oxobutyrate + CoA + oxidized methyl viologen	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis	propanoyl-CoA + CO2 + reduced methyl viologen	-	?
2-oxobutyrate + CoA + oxidized methyl viologen	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis DSM Z 2067	propanoyl-CoA + CO2 + reduced methyl viologen	-	?
indol-3 pyruvate + CoA + 2 oxidized methyl viologen	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis	?	-	?
indol-3 pyruvate + CoA + 2 oxidized methyl viologen	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis DSM Z 2067	?	-	?
oxaloacetate + CoA + oxidized methyl viologen	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis	?	-	?
oxaloacetate + CoA + oxidized methyl viologen	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis DSM Z 2067	?	-	?
pyruvate + CoA + 2 oxidized ferredoxin	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis	acetyl-CoA + CO2 + 2 reduced ferredoxin + 2 H+	-	?
pyruvate + CoA + 2 oxidized ferredoxin	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis DSM Z 2067	acetyl-CoA + CO2 + 2 reduced ferredoxin + 2 H+	-	?
pyruvate + CoA + 2 oxidized methyl viologen	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis	acetyl-CoA + CO2 + 2 reduced methyl viologen + 2 H+	-	?
pyruvate + CoA + 2 oxidized methyl viologen	the enzyme has a broad substrate specificity. In the presence of CoA, it oxidizes pyruvate, oxaloacetate, 2-oxobutyrate, and indol-3-pyruvate with specific activities of 7.4, 6.5, 3.6, and 0.3 U/mg, respectively. The enzyme reduces clostridial rubredoxin, clostridial and spinach ferredoxin, cytochrome c, FMN, and FAD. No activity is detected with NAD+, NADP+, and vitamin K1. The catalytic efficiencies or kcat/Km values for FAD and FMN are calculated to be 2400032000/min * M, which are about two orders of magnitude lower than observed for the likely physiological electron carriers of other pyruvate oxidoreductases. Therefore, it is unlikely that flavins are the physiological electron carrier of the methanococcal pyruvate oxidoreductase	Methanococcus maripaludis DSM Z 2067	acetyl-CoA + CO2 + 2 reduced methyl viologen + 2 H+	-	?

Subunits

Subunits	Comment	Organism
?	the low molecular weight enzyme contains five polypeptides: 47000 Da (alpha), 33000 Da (beta), 25000 (gamma) and 13000 Da (gamma). The subunit stoichiometry for the alpha:beta:gamma:delta subunits is 1:1.08:0.90:1.32. In addition it contains a fifth polypeptide (21500 Da)	Methanococcus maripaludis

Synonyms

Synonyms	Comment	Organism
POR	-	Methanococcus maripaludis

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Methanococcus maripaludis
60	-	at pH 7.3, the temperature optimum is 60°C	Methanococcus maripaludis

Temperature Range [°C]

Temperature Minimum [°C]	Temperature Maximum [°C]	Comment	Organism
37	60	the activity detected at 60°C is five times the activity detected at 37°C	Methanococcus maripaludis

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism
13	-	2-oxobutyrate	pH 8.6, 37°C	Methanococcus maripaludis
18	-	oxaloacetate	pH 8.6, 37°C	Methanococcus maripaludis
27	-	pyruvate	pH 8.6, 37°C	Methanococcus maripaludis

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.3	-	at 37 °C, maximal activity is obtained at pH 7.3	Methanococcus maripaludis
8.6	-	assay at	Methanococcus maripaludis

Cofactor

Cofactor	Comment	Organism
FAD	it is possible that flavins play an important regulatory or structural role in the enzyme	Methanococcus maripaludis
FMN	it is possible that flavins play an important regulatory or structural role in the enzyme	Methanococcus maripaludis
additional information	the enzyme is not coenzyme F420-dependent	Methanococcus maripaludis

General Information

General Information	Comment	Organism
physiological function	in autotrophic methanogens, pyruvate oxidoreductase plays a key role in the assimilation of CO2 and the biosynthesis of organic carbon	Methanococcus maripaludis

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism
3.6	-	2-oxobutyrate	pH 8.6, 37°C	Methanococcus maripaludis
68.2	-	oxaloacetate	pH 8.6, 37°C	Methanococcus maripaludis
234.8	-	pyruvate	pH 8.6, 37°C	Methanococcus maripaludis