Information on EC 1.12.98.1 - coenzyme F420 hydrogenase

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The expected taxonomic range for this enzyme is: Euryarchaeota

EC NUMBER
COMMENTARY hide
1.12.98.1
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RECOMMENDED NAME
GeneOntology No.
coenzyme F420 hydrogenase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
H2 + oxidized coenzyme F420 = reduced coenzyme F420
show the reaction diagram
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-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidation
-
-
-
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redox reaction
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-
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reduction
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
methanogenesis from CO2
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Methane metabolism
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Metabolic pathways
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Microbial metabolism in diverse environments
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SYSTEMATIC NAME
IUBMB Comments
hydrogen:coenzyme F420 oxidoreductase
An iron-sulfur flavoprotein (FAD) containing nickel. The enzyme from some sources contains selenocysteine. The enzyme also reduces the riboflavin analogue of F420, flavins and methylviologen, but to a lesser extent. The hydrogen acceptor coenzyme F420 is a deazaflavin derivative.
CAS REGISTRY NUMBER
COMMENTARY hide
9027-05-8
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain MF
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Manually annotated by BRENDA team
P80490: subunit beta, P80489: subunit alpha, P80491: subunit gamma
P80490 and P80489 and P80491
SwissProt
Manually annotated by BRENDA team
strain Fusaro, activity inducible by choice of substrate
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
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loss of F420H2 dehydrogenase, and therefore of the 420H2:heterodisulfide oxidoreductase system, does not measurably affect methanogenesis or growth in Methanosarcina barkeri
metabolism
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the preferred electron transport chain involves production of hydrogen gas in the cytoplasm, which then diffuses out of the cell, where it is reoxidized with transfer of electrons into the energy-conserving electron transport chain. This hydrogen-cycling metabolism leads directly to production of a proton motive force that can be used by the cell for ATP synthesis
physiological function
in the absence of hydrogenase Vhu, growth on hydrogen still occurs, albeit slowly
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
H2 + benzyl viologen
?
show the reaction diagram
H2 + coenzyme F0
reduced coenzyme F0
show the reaction diagram
H2 + coenzyme F420
reduced coenzyme F420
show the reaction diagram
H2 + methyl viologen
?
show the reaction diagram
H2 + oxidized benzyl viologen
reduced benzyl viologen
show the reaction diagram
H2 + oxidized coenzyme F0
reduced coenzyme F20
show the reaction diagram
H2 + oxidized coenzyme F420
reduced coenzyme F420
show the reaction diagram
H2 + oxidized methyl viologen
reduced methyl viologen
show the reaction diagram
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
H2 + coenzyme F420
reduced coenzyme F420
show the reaction diagram
H2 + oxidized coenzyme F420
reduced coenzyme F420
show the reaction diagram
H2 + oxidized methyl viologen
reduced methyl viologen
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
-
activating
KCl
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increases activity, maximal activity at 250 mM
Mg2+
-
activating
Nickel
selenium
sulfur
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
CO
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dead end inhibitor, competitive for H2, uncompetetitive versus F0
coenzyme F0
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substrate inhibition above 0.05 mM
methyl viologen
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noncompetitive for F0
Procion Blue HB
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inhibits F0 reduction
reduced coenzyme F0
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noncompetitive product inhibition
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
ammonium sulfate
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improves reactivation of enzyme with FAD after FAD depletion
KCl
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0.8 M activate F420-linked activity more than 10fold, slight inhibition of methyl viologen linked activity
Procion Blue HB
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stimulates methyl viologen reduction
additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.026 - 0.08
coenzyme F0
0.016 - 0.037
coenzyme F420
0.002 - 0.012
H2
0.42 - 1.56
methyl viologen
0.025
oxidized coenzyme F420
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37C, pH 7.5
0.0033
oxidized methyl viologen
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37C, pH 8.3
0.1
reduced coenzyme F0
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anaerobic conditions
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
725 - 2250
coenzyme F0
17.5 - 2050
coenzyme F420
725 - 4000
methyl viologen
353
oxidized coenzyme F420
Methanosarcina barkeri
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37C, pH 7.5
9226
oxidized methyl viologen
Methanosarcina barkeri
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37C, pH 8.3
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
25000 - 69000
H2
361
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0014
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methyl viologen-linked activity
0.0026
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methyl viologen-linked activity
0.032
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F420-linked activity
0.155
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F420-linked activity
0.34
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growth with H2 and CO2, enzyme activity depends on growth substrate
0.437
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growth on methanol, enzyme activity depends on growth substrate
0.847
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purified enzyme, F420-linked activity
1.24
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F420-linked activity
5.2
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purified enzyme with F420
10
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purified enzyme, F420-linked activity
11.1
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purified enzyme, benzyl viologen-linked activity
11.5
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37C, pH 7.5, substrate: oxidized coenzyme F420
16
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purified enzyme with methyl viologen
28
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C205P mutant enzyme with F420
49
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purified enzyme, F0-linked activity
50
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purified enzyme, methyl viologen-linked activity
63.2
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methyl viologen-linked activity
82.8
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37C, pH 7.2, cosubstrate: oxidized coenzyme F420
97.6
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purified enzyme with F420
98.6
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purified enzyme with F0
172
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purified enzyme with methyl viologen
345
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native enzyme with benzyl viologen
364
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native enzyme with F420
540
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C205P mutant enzyme with benzyl viologen
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 7.25
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deazaflavin reducing activity
6.5 - 7.5
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F420 reduction
7.5
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methyl viologen reduction
10
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optimum pH for methyl viologen reduction above pH 10
11
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optimum pH for methyl viologen reduction above pH 11
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5 - 8
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reverse reaction
5.5 - 10
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the methylviologen-reducing activity increased with pH from pH 5.5-10
7 - 11
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increasing methyl viologen activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25 - 90
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F420 reduction
40 - 103
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at 103C low activity with methyl viologen
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.3
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isoelectric focusing
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Methanothermobacter marburgensis (strain DSM 2133 / 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain DSM 2133 / 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain DSM 2133 / 14651 / NBRC 100331 / OCM 82 / Marburg)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
102000
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smallest active species, gel filtration
105000
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smallest active species, gel filtration, sucrose density centrifugation
109000
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gel filtration
115000
150000
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gel filtration
170000
198000
340000
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gel filtration, enzyme tends to aggregate, aggregated form MW 1300000
380000
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gel filtration
500000
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native enzyme bigger than 500000
600000
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gel filtration
670000
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membrane-associated protein complex, native PAGE
745000
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gel filtration, sucrose density centrifugation
790000
800000
845000
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non-denaturing PAGE, large enzyme form
990000
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native SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexamer
multimer
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
structure determined by near-atomic resolution cryo-electron microscopy with and without bound substrate F420. The 1.2-MDa complex contains 12 copies of the heterotrimer, which form a spherical protein shell with a hollow core. There is strong electron density of the chains of metal clusters running parallel to the protein shell, and the F420-binding site is located at the end of the chain near the outside of the spherical structure
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
10
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unstable above pH 10
395095
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0 - 5
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reactivated enzyme stable
25 - 30
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reactivated enzyme unstable
65
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methyl viologen-reducing activity heat resistent up to 65C, F420-reducing activity 75% inactivated at 65C
70
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stable for more than 3 h, methyl viologen-linked activity more stable against heating than F420-linked activity
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
0.03 mM FAD stabilizes enzyme
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FAD stabilizes F420-linked activity against heat inactivation, generally stabilizing
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freezing and thawing changes properties of enzyme, different behavior in anion exchange chromatography
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unstable in prereduced buffer, dramatic decline of activity after 2 h
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OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
aerobic purification yields the stable but inactive enzyme, can be reactivated under reducing conditions
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395086
enzyme activity is unstable under reducing conditions
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727399
extremely oxygen sensitive, can be reactivated by incubation with molecular hydrogen and dithiothreitol
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395079
more stable under aerobic conditions, reversible loss of activity with oxygen can be restored with dithionite
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395082
purified enzyme requires reductive reactivation before assay, reactivation possible with H2 or glucose and glucose oxidase
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395093
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 1 year, inactive form maintains 70% of initial activity after reactivation
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-70C, loss of high temperature activity
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4C, 4 weeks, 25% loss of activity towards F420, 97% loss of activity towards methyl viologen
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4C, 5% loss of activity within 24 h
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liquid nitrogen, in presence of 50% sucrose and 10% glycerol, stable
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room temperature, activated enzyme loses 20% of inital activity within 24 h
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli
P19496 and P19499 and P19498
expression in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C205P
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no effect on benzyl viologen reduction, 10 fold decrease in F420 reduction, wild type enzyme contains 3[4Fe-4S], mutant enzyme contains 1[3Fe-4S]
additional information
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Methanosarcina barkeri Delta fpo mutants: strain WWM85 Delta hpt::PmcrB-Phi C31int-attP, strain WWM86 Delta hpt::PmcrB-Phi C31int-attB, strain WWM71 Delta hpt::PmcrB-Phi C31int-attB, Delta fpoA-O Deletion of fpo by markerless exchange with pDK4 in WWM86, strain WWM123 Delta hpt::PmcrB-Phi C31int-attP, Delta fpoF Deletion of fpoF by markerless exchange with pDK13 in WWM85, strain WWM116 Delta hpt::PmcrB-Phi C31int-attP, Delta freAEGB Deletion of fre by markerless exchange with pGK6 in WWM85, strain WWM122 Delta hpt::PmcrB-Phi C31int-attB, Delta frhADGB::pac-hpt Deletion of frh with ApaI/NotI-digested pAMG81 in WWM86, strain WWM108 Delta hpt::PmcrB-Phi C31int-attB, Delta fpoA-O, Delta frhADGB::pac-hpt Deletion of frh with ApaI/NotI-digested pAMG81 in WWM71, strain WWM145 Delta hpt::PmcrB-Delta C31int-attP, Delta fpoF, Delta frhADGB::pac-hpt Deletion of frh with ApaI/NotI-digested pAMG81 in WWM123
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