Information on EC 2.8.4.1 - coenzyme-B sulfoethylthiotransferase

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The expected taxonomic range for this enzyme is: Archaea, Bacteria

EC NUMBER
COMMENTARY hide
2.8.4.1
-
RECOMMENDED NAME
GeneOntology No.
coenzyme-B sulfoethylthiotransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
methyl-CoM + CoB = CoM-S-S-CoB + methane
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
sulfo ethyl group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
-
-
Methane metabolism
-
-
methyl-coenzyme M reduction to methane
-
-
Microbial metabolism in diverse environments
-
-
coenzyme M biosynthesis
-
-
SYSTEMATIC NAME
IUBMB Comments
methyl-CoM:CoB S-(2-sulfoethyl)thiotransferase
This enzyme catalyses the final step in methanogenesis, the biological production of methane. This important anaerobic process is carried out only by methanogenic archaea. The enzyme can also function in reverse, for anaerobic oxidation of methane. The enzyme requires the hydroporphinoid nickel complex coenzyme F430. Highly specific for coenzyme B with a heptanoyl chain; ethyl CoM and difluoromethyl CoM are poor substrates. The sulfide sulfur can be replaced by selenium but not by oxygen.
CAS REGISTRY NUMBER
COMMENTARY hide
53060-41-6
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
M.o.H. and M.o.H.G.
-
-
Manually annotated by BRENDA team
Methanocellales sp.
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
isozyme MCR I; isozymes MCR I, gene mcrA
-
-
Manually annotated by BRENDA team
Methanogenic bacterium
from bovine rumen fluid
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
Q49605: alpha subunit, Q49601: beta subunit, Q49604: gamma subunit
Q49605 and Q49601 and Q49604
SwissProt
Manually annotated by BRENDA team
DSM 2139
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
P07962: alpha subunit, P07955: beta subunit, P07964: gamma subunit
P07962 and P07955 and P07964
SwissProt
Manually annotated by BRENDA team
; formerly Methanosarcina frisia
-
-
Manually annotated by BRENDA team
formerly Methanosarcina frisia
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
formerly Methanobacterium thermoautotrophicum, strain Marburg
-
-
Manually annotated by BRENDA team
formerly Methanobacterium wolfei
-
-
Manually annotated by BRENDA team
methanotrophic archaeon
-
-
-
Manually annotated by BRENDA team
no activity in Methanobrevibacter ruminantium
-
-
-
Manually annotated by BRENDA team
no activity in Methanobrevibacter ruminantium M-1
-
-
-
Manually annotated by BRENDA team
subunits alpha, beta and gamma; consortia of microorganisms collected from a Black Sea mat (ANME-1)
D1JBK2 and D1JBK3 and D1JBK4
UniProt
Manually annotated by BRENDA team
the methanogenic community in hydrothermally active sediments of Guaymas Basin (Gulf of California, Mexico) is analyzed by PCR amplification, cloning, and sequencing of methyl coenzyme M reductase genes. Members of the Methanomicrobiales and Methanosarcinales dominate the mcrA clone libraries from the upper 15 cm of the sediments. Within the H2 /CO2 - and formate-utilizing family Methanomicrobiales, two mcrA and 16S rRNA lineages are closely affiliated with cultured species of the genera Methanoculleus and Methanocorpusculum. The most frequently recovered mcrA PCR amplicons within the Methanomicrobiales does not branch with any cultured genera. Within the nutritionally versatile family Methanosarcinales, one 16S rRNA amplicon and most of the mcrA PCR amplicons are affiliated with the obligately acetate utilizing species Methanosaeta concilii. The mcrA clone libraries also includes phylotypes related to the methyl-disproportionating genus Methanococcoides. Two mcrA and two 16S rRNA lineages within the Methanosarcinales are unrelated to any cultured genus
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2-(methylthio)ethanesulfonate + N-(7-mercaptoheptanoyl)threonine 3-O-phosphate
CoM-S-S-CoB + methane
show the reaction diagram
2-(methylthio)ethansulfonate + N-(7-mercaptoheptanoyl)threonine 3-O-phosphate
CoM-S-S-CoB + methane
show the reaction diagram
-
i.e. CoM and CoB
-
-
?
CH3-S-CoM + HS-CoB
CoM-S-S-CoB + methane
show the reaction diagram
CH3-S-CoM + HS-CoB6
CoM-S-S-CoB6 + methane
show the reaction diagram
-
i.e. N-7-mercaptohexanoylthreonine phosphate
-
-
?
CH3-S-CoM + HS-CoB8
CoM-S-S-CoB8 + methane
show the reaction diagram
-
a two-electron transfer reaction
-
-
?
CH3-S-CoM + HS-CoB9
CoM-S-S-CoB9 + methane
show the reaction diagram
-
-
-
-
?
CH3-S-CoM + SH-CoB
CoM-S-S-CoB + methane
show the reaction diagram
CH3-S-CoM + SH-CoB5
CoM-S-S-CoB5 + methane
show the reaction diagram
CH3-S-CoM + SH-CoB6
CoM-S-S-CoB6 + methane
show the reaction diagram
CH3-S-CoM + SH-CoB8
CoM-S-S-CoB8 + methane
show the reaction diagram
CH3-S-CoM + SH-CoB9
CoM-S-S-CoB9 + methane
show the reaction diagram
CH3-S-CoM3 + HS-CoB8
CoM3-S-S-CoB8 + methane
show the reaction diagram
-
-
-
-
?
CoM-S-S-CoB + methane
methyl-CoM + CoB
show the reaction diagram
-
-
-
-
?
ethyl coenzyme M + coenzyme B
ethane + CoM-S-S-CoB
show the reaction diagram
-
1% of the activity with methyl coenzyme M
-
-
?
ethyl-CoM + CoB
CoM-S-S-CoB + ethane
show the reaction diagram
-
-
-
-
?
methyl coenzyme M + coenzyme B
methane + CoM-S-S-CoB
show the reaction diagram
methyl-coenzyme M + coenzyme B
methane + CoM-S-S-CoB
show the reaction diagram
methyl-coenzyme M + N-(7-mercaptoheptanoyl)threonine 3-O-phosphate (coenzyme B)
methane + CoB-S-S-CoM
show the reaction diagram
-
i.e. methyl-SCoM
a the mixed disulfide
-
?
methyl-coenzyme M + N-(7-mercaptoheptanoyl)threonine 3-O-phosphate (coenzyme B)
methane + CoM-S-S-CoB
show the reaction diagram
methyl-CoM + CoB
CoM-S-S-CoB + methane
show the reaction diagram
methylmercaptopropionate + HS-CoB
?
show the reaction diagram
-
is about 110fold less reactive than the natural substrate methyl-SCoM
-
-
?
additional information
?
-
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
2-(methylthio)ethanesulfonate + N-(7-mercaptoheptanoyl)threonine 3-O-phosphate
CoM-S-S-CoB + methane
show the reaction diagram
-
i.e. CoM and CoB
-
-
?
2-(methylthio)ethansulfonate + N-(7-mercaptoheptanoyl)threonine 3-O-phosphate
CoM-S-S-CoB + methane
show the reaction diagram
-
i.e. CoM and CoB
-
-
?
CH3-S-CoM + HS-CoB
CoM-S-S-CoB + methane
show the reaction diagram
CH3-S-CoM + SH-CoB
CoM-S-S-CoB + methane
show the reaction diagram
methyl coenzyme M + coenzyme B
methane + CoM-S-S-CoB
show the reaction diagram
methyl-coenzyme M + N-(7-mercaptoheptanoyl)threonine 3-O-phosphate (coenzyme B)
methane + CoB-S-S-CoM
show the reaction diagram
-
i.e. methyl-SCoM
a the mixed disulfide
-
?
methyl-coenzyme M + N-(7-mercaptoheptanoyl)threonine 3-O-phosphate (coenzyme B)
methane + CoM-S-S-CoB
show the reaction diagram
methyl-CoM + CoB
CoM-S-S-CoB + methane
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
coenzyme F430
additional information
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ni(2+)
Nickel
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(+)-(2S,3R)-N-[7-(methylthio)heptanoyl]-O-phospho-L-threonine
-
CoB substrate thioester derivative, synthesis, spectral analysis and binding structure, overview
(+)-(2S,3R)-N-[7-(trifluoromethylthio)heptanoyl]-O-phospho-L-threonine
-
fluorescent CoB substrate thioester derivative, synthesis, spectral analysis and binding structure, overview
1-propanesulfonate
-
-
2-azidoethanesulfonate
-
competitive, reversible
2-bromoethanesulfonate
3-azidopropanesulfonate
-
-
3-bromopropane sulfonate
3-bromopropanesulfonate
3-Bromopropionate
3-chloropropanesulfonyl chloride
-
-
3-fluoropropanesulfonate
-
-
3-hydroxypropanesulfonate
-
-
3-iodopropane sulfonate
-
-
3-iodopropanesulfonate
-
-
3-mercapto-1-propanesulfonate
-
-
4-bromobutanesulfonate
4-bromobutyrate sulfonate
-
when reacted with 4-bromobutyrate, MCRred1 forms the alkyl-Ni(III) MCRXA state and then self-reactivation to regenerate the Ni(I) MCRred1 state and a bromocarboxy ester
-
7-bromoheptanoylthreonine phosphate
-
substrate analogue of coenzyme B in which the thiol group is substituted by bromine, potent inhibitor, competitive with respect to methyl-coenzyme M and B
allyl-coenzyme M
-
reversible
bromoethane sulfonate
D1JBK2 and D1JBK3 and D1JBK4
-
bromopropanesulfonate
-
BPS, a potent inhibitor and reversible redox inactivator that reacts with MCRred1 to form an EPR-active state called MCRPS, which is an alkyl-nickel species. Treatment of MCRPS with free thiol containing compounds leads to reconvertion to the active MCRred1 state
chloromethanesulfonate
-
-
CO
-
little blocking effect on the hydrogenase function of component A
coenzyme M
CoM-S-S-CoB
-
50% inhibition at 0.6 mM
cyano-coenzyme M
-
irreversible
ethanol
-
concentrations above 0.005 ml/0.2 ml assay
HS-CoM
-
the unmethylated coenzyme M is a reversible competitive inhibitor
N-6-mercaptohexanoylthreonine phosphate
-
competitive with coenzyme B
N-7(methylthio)-heptanoylthreonine phosphate
-
competitive with coenzyme B
N-8-mercaptooctanoylthreonine phosphate
-
competitive with coenzyme B
O-phosphono-N-(5-sulfanylpentanoyl)-L-threonine
P11558 and P11560 and P11562
-
O-phosphono-N-(6-sulfanylhexanoyl)-L-threonine
P11558 and P11560 and P11562
-
O2
-
total loss of activity of hydrogenase activity of component A
propyl-coenzyme M
-
reversible
seleno-coenzyme M
-
irreversible
trifluoromethyl coenzyme M
-
irreversible
-
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
ADP
-
in absence of reducing agents
arsenate
-
in absence of reducing agents
Cobalamin
coenzyme F-420
-
enzyme system is stimulated
EDTA
-
in absence of reducing agents
H2
-
suggested to be involved in the activation of the methylreductase, preincubation
phosphate
-
in absence of reducing agents
reduced ferredoxin
-
reactivation after purification
sulfate
-
in absence of reducing agents
Ti(III) citrate
vitamin B12
-
enzyme system is stimulated
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.03 - 3.5
CH3-S-CoM
62
CH3-S-CoM3
-
pH 10.0, 25C
0.169
CoB
-
at pH 7.6 and 25C
0.023 - 0.6
coenzyme B
20
ethyl coenzyme M
-
-
0.016
HS-CoB8
-
pH 10.0, 25C
0.46
HS-CoB9
-
pH 10.0, 25C
0.0033 - 5
methyl coenzyme M
2
methyl-coenzyme M
-
-
0.34
SH-CoB5
-
pH not specified in the publication, at 25C
0.08
SH-CoB6
-
pH not specified in the publication, at 25C
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
18
CoB
Methanothermobacter marburgensis
-
at pH 7.6 and 25C
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.001
2-azidoethanesulfonate
-
-
0.004 - 0.1
2-bromoethanesulfonate
0.04
3-azidopropanesulfonate
-
-
0.00005
3-bromopropane sulfonate
-
-
0.00005 - 0.0001
3-bromopropanesulfonate
0.05
3-fluoropropanesulfonate
-
below
4
3-hydroxypropanesulfonate
-
-
0.001
3-iodopropanesulfonate
-
below
0.006
4-bromobutanesulfonate
-
-
0.005
7-bromoheptanoylthreonine phosphate
-
-
2
allyl-coenzyme M
-
-
0.25
chloromethanesulfonate
-
-
0.0001
N-6-mercaptohexanoylthreonine phosphate
-
competitive with coenzyme B
0.0001
O-phosphono-N-(6-sulfanylhexanoyl)-L-threonine
P11558 and P11560 and P11562
pH and temperature not specified in the publication
5
propyl-coenzyme M
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0004
2-bromoethanesulfonate
Methanobrevibacter ruminantium
-
pH and temperature not specified in the publication
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.006
-
pH and temperature not specified in the publication
0.5 - 2
-
in presence of dithiothreitol and of reduced corrinoids or Ti(III) citrate
1 - 2
-
in presence of 15 mM dithiothreitol and 0.3 mM aquocobalamin or 0.15 mM Ti(III) citrate
2.5
-
purified, addition of Ti(III) citrate, dithiothreitol and cobalamin
3
-
extract of H2-reduced cells
20.1
-
purification from H2-reduced cells
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
-
60 nmol of methane are formed per minute per mg protein
additional information
-
pH 10.0 is optimal for activation of MCRox1 to MCRred1 with Ti(III) citrate
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
-
growth temperature 65C
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
-
no activity found in membrane fraction
-
Manually annotated by BRENDA team
PDB
SCOP
CATH
ORGANISM
UNIPROT
Methanosarcina barkeri (strain Fusaro / DSM 804)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
Methanothermobacter marburgensis (strain ATCC BAA-927 / DSM 2133 / JCM 14651 / NBRC 100331 / OCM 82 / Marburg)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
28700
-
x * 59800 + x * 47100 + x * 28700, SDS-PAGE
30400
-
1 * 63900, 1 * 41700, 1 * 30400, SDS-PAGE
33000
-
1 * 69000, 1 * 42000 and 1 * 33000, alpha, beta and gamma
37000
-
2 * 66000, 2 * 48000, 2 * 37000, SDS-PAGE
41700
-
1 * 63900, 1 * 41700, 1 * 30400, SDS-PAGE
42000
-
1 * 69000, 1 * 42000 and 1 * 33000, alpha, beta and gamma
47100
-
x * 59800 + x * 47100 + x * 28700, SDS-PAGE
48000
-
2 * 66000, 2 * 48000, 2 * 37000, SDS-PAGE
59800
-
x * 59800 + x * 47100 + x * 28700, SDS-PAGE
63900
-
1 * 63900, 1 * 41700, 1 * 30400, SDS-PAGE
66000
-
2 * 66000, 2 * 48000, 2 * 37000, SDS-PAGE
69000
-
1 * 69000, 1 * 42000 and 1 * 33000, alpha, beta and gamma
132000
-
nondenaturing slab gel electrophoresis
141000
-
gel filtration
270000
-
gel filtration
280000
300000
additional information
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 59800 + x * 47100 + x * 28700, SDS-PAGE
dimer of heterotrimers
-
-
heterohexamer
heterotrimer
hexamer
trimer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
side-chain modification
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, crystals of the inactive enzyme are obtained with PEG 550 monomethyl ether as precipitant. Diffraction data to 2.7 A resolution are collected from one crystal of methyl-coenzyme M reductase from Methanopyrus kandleri with a completeness of 63%. Due to the low completeness of the data, refinement of the structure is only possible constraining the 2-fold non-crystallographic symmetry of the methyl-coenzyme M reductase molecule. Comparison of crystal structures of methyl-coenzyme M reductase from Methanosarcina barkeri (growth temperature optimum, 37C), Methanopyrus kandleri (growth temperature optimum, 98C) and Methanobacterium thermoautotrophicum (growth temperature optimum, 65C); the crystal structures of methyl-coenzyme M reductase from Methanosarcina barkeri and Methanopyrus kandleri are determined and compared with the known structure of MCR from Methanobacterium thermoautotrophicum. The active sites of enzyme from Methanosarcina barkeri and Methanopyrus kandleri are almost identical to that of Methanobacterium thermoautotrophicum and predominantly occupied by coenzyme M and coenzyme B. Crystals of the inactive enzyme from Methanopyrus kandleri are obtained by hanging drop method with PEG 550 monomethylether as precipitant
Q49605 and Q49601 and Q49604
hanging drop method, comparison of crystal structures of methyl-coenzyme M reductase from Methanosarcina barkeri (growth temperature optimum, 37C), Methanopyrus kandleri (growth temperature optimum, 98C) and Methanobacterium thermoautotrophicum (growth temperature optimum, 65C); the crystal structures of methyl-coenzyme M reductase from Methanosarcina barkeri and Methanopyrus kandleri are determined and compared with the known structure of MCR from Methanobacterium thermoautotrophicum. The active sites of enzyme from Methanosarcina barkeri and Methanopyrus kandleri are almost identical to that of Methanobacterium thermoautotrophicum and predominantly occupied by coenzyme M and coenzyme B. The electron density at 1.6 A resolution of the Methanosarcina barkeri enzyme reveals that four of the modified amino acid residues of enzyme from Methanopyrus thermoautotrophicum, namely a thiopeptide, an S-methylcysteine, a 1-N-methylhistidine and a 5-methylarginine are also present. Crystals of the enzyme from Methanosarcina barkeri are grown using a reservoir condition with PEG 5000 monomethylether as precipitant and glycerol as cryoprotectant
P07962 and P07955 and P07964
enzyme alone and in complex with substrates, sitting drop vapor diffusion method, using 100 mM Na-HEPES, pH 7.3-8.0, 150 mM magnesium acetate, and 20-22% (w/v) PEG 400
P11558 and P11560 and P11562
hanging drop vapor diffusion method, crystal form M obtained with 2-methyl-2,4-pentanediol grown within two months, form P grows from polyethylene glycol 400 within two weeks at 4C, both crystal forms have one molecule per assymetric unit
-
sitting drop vapor diffusion method, using 25% (w/v) PEG 400, 0.1M Tris pH 8.5 and 0.2 M Li2SO4
D1JBK2 and D1JBK3 and D1JBK4
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
9
-
above this pH value and 60C isoenzyme I is slowly denatured
393247
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4
-
11 h, fully active
23
-
N2 atmosphere, 2 h, 28% loss of original activity
60
-
pH above 9.0 isoenzyme I is slowly denatured
95
-
10 min under anaerobic conditions, heat-labile components A and C lose activity
121
-
1-2 h, heat-stable component B, no loss of activity
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
activity is stabilized by high concentrations of ammonium sulfate
-
methyl-coenzyme M stabilizes enzyme activity
-
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
high concentrations of both H2 and CO2 favour the expression of isoenzyme II, low concentrations favour the expression of isoenzyme I
-
393248
oxygen causes irreversible loss of methane forming activity, component C is oxygen-stable
-
393259
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 50 mM potassium PIPES buffer, pH 6.2, 15 mM MgCl2, 10 mM 2-mercaptoethanol
-
-20C, stored under N2 atmosphere, 50 mM anaerobic Tris-HCl, pH 7.6, 150 mM NaCl, 1 mM Ti(III) citrate
-
-70C, rapidly frozen, yellow precipitate, suspended in 100%-saturated ammonium sulfate, stored unter N2, remains active
-
-80C, under N2 atmosphere, 50 mM PIPES, pH 7.2, 40% v/v glycerol, placed in H2 gas phase, stable for weeks
-
0C, enzyme preparation, 100% H2
-
4C, 11 h, fully active
-
4C, component C, 96% homogeneous, under N2 atmosphere, several months without any loss of activity in multicomponent assay
-
4C, pH 7.0, 4 h, 50% loss of activity of red1 form
-
4C, under N2 atmosphere, 50 mM potassium phosphate, pH 6.7, 50% decrease of specific acitivity within 12 hours
-
ice bath temperature, hydrogenase stable 3-4 weeks
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liquid N2, 50 mM potassium phosphate buffer, pH 7.0, 1 M potassium acetate, 10 mM 2-mercaptoethanol, 10% w/v glycerol
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storage in frozen stage not possible, thawing leads to a rapid loss of activity
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three freeze-thaw cycles, fully active
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
4C, strictly anaerobic conditions, fractional ammonium-sulfate precipitation; copurification of isoenzymes, separation of isoenzymes by anion-exchange chromatography
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active native enzyme to homogeneity
-
active native isozyme MCR I in presence of HS-CoM
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active native MCR; native enzyme, during purification the enzyme lost its MCR-red2 signal owing to the removal of HS-CoB
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all buffers that are used during purification process are supplemented with stabilising methyl coenzyme M, further chromatography steps with phenyl-superose, Mono-Q and Sephadex G25 result in 90% loss of specific activity; under 5% H2/95% N2 condition, fractionated ammonium sulfate precipitation
-
anaerobic conditions, ion-exchange column, gel-filtration
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anaerobic, different components purified seperately, component C ion-exchange chromatography, hydrophobic interaction chromatography, component A2 affinity chromatography, ion-exchange chromatography, component A1 and A3 are obtained by gel filtration, hydrophobic interaction chromatography
-
copurification of isoenzymes, separation of isoenzymes by anion-exchange chromatography; hydrophobic interaction chromatography, Sepharose beads which contain antibodies specific for each isoenzyme, immunoadsorption method; under 5% H2/95% N2 condition, fractionated ammonium sulfate precipitation
-
in the silent, red1c, or ox1 states
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membrane filtration, on Mono Q, Superose-6 and Phenyl-Superose column
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purification of native MCRox1, and of native MCRred1, the latter by ammonium sulfate fractionation and ion exchange chromatography
-
Q-Sepharose, Phenyl-Superose HR 5/5, Superose 6 HR 10/30 column
-
sucrose density gradient cetrifugation, Q-Sepharose, Phenyl-Superose 5/5 column, anaerobic
-
under 5% H2/95% N2 condition, fractionated ammonium sulfate precipitation
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression of two ORF gene products in Escherichia coli DS 410, expressed proteins are suggested to be important for enzyme activity, but neither stimulatory nor inhibitory effects of these gene products are determined
-
genes in operons mrt and mcr, DNA and amino acid sequence determination, promoter region, determination of operator regions of the mcr and mrt operons, and of cis elements and trans-acting factors responsible for the gene expression of MCRs by using electrophoretic mobility shift assay and affinity particle purification. IMP dehydrogenase-related protein VII, IMPDH VII encoded by MTH126, is a plausible candidate for the transcriptional regulator of the mcr operon in this methanogen, the binding site of IMPDH VII mostly overlaps the factor B-responsible element-TATA box of the mcr operon, overview. Expression in Escherichia coli strain BL21(DE3)
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
environmental protection
-
expression of methyl-coenzyme M reductase from an unculturable organism in Methanosarcina acetivorans to effectively run methanogenesis in reverse. Methanosarcina acetivorans cells heterologously producing methyl-coenzyme M reductase consume up to 9% of methane (corresponding to 109 micromol of methane) after 6 weeks of anaerobic growth on methane and utilize 10 mM FeCl3 as an electron acceptor. When incubated on methane for 5 days, high-densities of cells consume 15% methane (corresponding to 143 micromol of methane), and produce 10.3 mM acetate (corresponding to 52 micromol of acetate)
synthesis
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expression of methyl-coenzyme M reductase from an unculturable organism in Methanosarcina acetivorans to effectively run methanogenesis in reverse. Methanosarcina acetivorans cells heterologously producing methyl-coenzyme M reductase consume up to 9% of methane (corresponding to 109 12 micromol of methane) after 6 weeks of anaerobic growth on methane and utilize 10 mM FeCl3 as an electron acceptor. When incubated on methane for 5 days, high-densities of cells consume 15% methane (corresponding to 143 16 micromol of methane), and produce 10.3 mM acetate (corresponding to 52 4 mol of acetate)
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