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
coenzyme M biosynthesis
-
-
Metabolic pathways
-
-
Methane metabolism
-
-
methyl-coenzyme M reduction to methane
-
-
Microbial metabolism in diverse environments
-
-
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
M.o.H. and M.o.H.G.
-
-
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
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
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
Methanothermobacter thermautotrophicum
Methanothermobacter thermautotrophicum DELTAH
deltaH
-
-
Manually annotated by BRENDA team
Methanothermobacter thermautotrophicum Marburg
formerly Methanobacterium wolfei
-
-
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
i.e. anaerobic methanotrophic archaeal population 1, ANME-1
-
-
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
-
-
?
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
Mg2+
Methanothermobacter thermautotrophicum
-
-
Ni(2+)
Ni3+
-
the enzyme contains nickel, Ni(III)-methyl is an intermediate in methane formation
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
Methanothermobacter thermautotrophicum
-
competitive, reversible
2-bromoethanesulfonate
3-azidopropanesulfonate
Methanothermobacter thermautotrophicum
-
-
3-bromopropane sulfonate
3-bromopropanesulfonate
3-Bromopropionate
3-chloropropanesulfonyl chloride
-
-
3-fluoropropanesulfonate
Methanothermobacter thermautotrophicum
-
-
3-hydroxypropanesulfonate
Methanothermobacter thermautotrophicum
-
-
3-iodopropane sulfonate
-
-
3-iodopropanesulfonate
Methanothermobacter thermautotrophicum
-
-
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
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
-
CO
Methanothermobacter thermautotrophicum
-
little blocking effect on the hydrogenase function of component A
coenzyme M
CoM-S-S-CoB
Methanothermobacter thermautotrophicum
-
50% inhibition at 0.6 mM
cyano-coenzyme M
-
irreversible
ethanol
Methanothermobacter thermautotrophicum
-
concentrations above 0.005 ml/0.2 ml assay
HS-CoM
-
the unmethylated coenzyme M is a reversible competitive inhibitor
N-6-mercaptohexanoylthreonine phosphate
Methanothermobacter thermautotrophicum
-
competitive with coenzyme B
N-7(methylthio)-heptanoylthreonine phosphate
Methanothermobacter thermautotrophicum
-
competitive with coenzyme B
N-8-mercaptooctanoylthreonine phosphate
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
in absence of reducing agents
arsenate
Methanothermobacter thermautotrophicum
-
in absence of reducing agents
CN-B12
Methanothermobacter thermautotrophicum
-
-
Cobalamin
coenzyme F-420
Methanothermobacter thermautotrophicum
-
enzyme system is stimulated
dithiothreitol
Methanothermobacter thermautotrophicum
-
-
EDTA
Methanothermobacter thermautotrophicum
-
in absence of reducing agents
H2
Methanothermobacter thermautotrophicum
-
suggested to be involved in the activation of the methylreductase, preincubation
phosphate
Methanothermobacter thermautotrophicum
-
in absence of reducing agents
reduced corrinoids
Methanothermobacter thermautotrophicum
-
-
-
reduced ferredoxin
-
reactivation after purification
sulfate
Methanothermobacter thermautotrophicum
-
in absence of reducing agents
Ti(III) citrate
vitamin B12
Methanothermobacter thermautotrophicum
-
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
0.023 - 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
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.001
2-azidoethanesulfonate
Methanothermobacter thermautotrophicum
-
-
0.004 - 0.1
2-bromoethanesulfonate
0.04
3-azidopropanesulfonate
Methanothermobacter thermautotrophicum
-
-
0.00005
3-bromopropane sulfonate
0.0001
3-bromopropanesulfonate
Methanothermobacter thermautotrophicum
-
for both isoenzymes
0.05
3-fluoropropanesulfonate
Methanothermobacter thermautotrophicum
-
below
4
3-hydroxypropanesulfonate
Methanothermobacter thermautotrophicum
-
-
0.001
3-iodopropanesulfonate
Methanothermobacter thermautotrophicum
-
below
0.006
4-bromobutanesulfonate
Methanothermobacter thermautotrophicum
-
-
0.005
7-bromoheptanoylthreonine phosphate
Methanothermobacter thermautotrophicum
-
-
2
allyl-coenzyme M
-
-
0.25
chloromethanesulfonate
Methanothermobacter thermautotrophicum
-
-
0.0001
N-6-mercaptohexanoylthreonine phosphate
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
0.00054
Methanothermobacter thermautotrophicum
-
-
0.006
-
pH and temperature not specified in the publication
0.075
Methanothermobacter thermautotrophicum
-
up tp
0.1
Methanothermobacter thermautotrophicum
-
-
0.5 - 2
Methanothermobacter thermautotrophicum
-
in presence of dithiothreitol and of reduced corrinoids or Ti(III) citrate
0.72
Methanothermobacter thermautotrophicum
-
CO2-oxidized cells
1 - 2
Methanothermobacter thermautotrophicum
-
in presence of 15 mM dithiothreitol and 0.3 mM aquocobalamin or 0.15 mM Ti(III) citrate
1.9
Methanothermobacter thermautotrophicum
-
for component C
2.5
Methanothermobacter thermautotrophicum
-
purified, addition of Ti(III) citrate, dithiothreitol and cobalamin
3
Methanothermobacter thermautotrophicum
-
extract of H2-reduced cells
4.5
Methanothermobacter thermautotrophicum
-
isoenzyme I
11.7
Methanothermobacter thermautotrophicum
-
isoenzyme II
20.1
Methanothermobacter thermautotrophicum
-
purification from H2-reduced cells
100
Methanothermobacter thermautotrophicum
-
-
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6
Methanothermobacter thermautotrophicum
-
assay at
6.7
Methanothermobacter thermautotrophicum
-
assay at
7
-
60 nmol of methane are formed per minute per mg protein
7 - 7.2
Methanothermobacter thermautotrophicum
-
assay at
7 - 7.5
Methanothermobacter thermautotrophicum
-
isoenzyme I
7.5 - 8
Methanothermobacter thermautotrophicum
-
isoenzyme II
additional information
-
pH 10.0 is optimal for activation of MCRox1 to MCRred1 with Ti(III) citrate
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
62
Methanothermobacter thermautotrophicum
-
assay
65
Methanothermobacter thermautotrophicum
-
assay at
68 - 70
Methanothermobacter thermautotrophicum
-
both isoenzymes
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 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)
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)
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)
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)
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
132000
-
nondenaturing slab gel electrophoresis
141000
-
gel filtration
150000
Methanothermobacter thermautotrophicum
-
PAGE
280000
300000
additional information
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 59800 + x * 47100 + x * 28700, SDS-PAGE
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
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
pH above 9.0 isoenzyme I is slowly denatured
95
Methanothermobacter thermautotrophicum
-
10 min under anaerobic conditions, heat-labile components A and C lose activity
121
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
methyl-coenzyme M stabilizes enzyme activity
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
393248
oxygen causes irreversible loss of methane forming activity, component C is oxygen-stable
Methanothermobacter thermautotrophicum
-
393259
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 50 mM potassium PIPES buffer, pH 6.2, 15 mM MgCl2, 10 mM 2-mercaptoethanol
Methanothermobacter thermautotrophicum
-
-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
Methanothermobacter thermautotrophicum
-
-80C, under N2 atmosphere, 50 mM PIPES, pH 7.2, 40% v/v glycerol, placed in H2 gas phase, stable for weeks
Methanothermobacter thermautotrophicum
-
0C, enzyme preparation, 100% H2
Methanothermobacter thermautotrophicum
-
4C, 11 h, fully active
-
4C, component C, 96% homogeneous, under N2 atmosphere, several months without any loss of activity in multicomponent assay
Methanothermobacter thermautotrophicum
-
4C, pH 7.0, 4 h, 50% loss of activity of red1 form
Methanothermobacter thermautotrophicum
-
4C, under N2 atmosphere, 50 mM potassium phosphate, pH 6.7, 50% decrease of specific acitivity within 12 hours
Methanothermobacter thermautotrophicum
-
ice bath temperature, hydrogenase stable 3-4 weeks
Methanothermobacter thermautotrophicum
-
liquid N2, 50 mM potassium phosphate buffer, pH 7.0, 1 M potassium acetate, 10 mM 2-mercaptoethanol, 10% w/v glycerol
-
storage in frozen stage not possible, thawing leads to a rapid loss of activity
Methanothermobacter thermautotrophicum
-
three freeze-thaw cycles, fully active
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
4C, strictly anaerobic conditions, fractional ammonium-sulfate precipitation; copurification of isoenzymes, separation of isoenzymes by anion-exchange chromatography
Methanothermobacter thermautotrophicum
-
active native enzyme to homogeneity
-
active native isozyme MCR I in presence of HS-CoM
-
active native MCR; native enzyme, during purification the enzyme lost its MCR-red2 signal owing to the removal of HS-CoB
-
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
Methanothermobacter thermautotrophicum
-
anaerobic conditions
Methanothermobacter thermautotrophicum
-
anaerobic conditions, ion-exchange column, gel-filtration
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
in the silent, red1c, or ox1 states
-
membrane filtration, on Mono Q, Superose-6 and Phenyl-Superose column
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
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
Methanothermobacter thermautotrophicum
-
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)
Methanothermobacter thermautotrophicum
-
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
-
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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