Information on EC 2.1.1.5 - betaine-homocysteine S-methyltransferase

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

EC NUMBER
COMMENTARY hide
2.1.1.5
-
RECOMMENDED NAME
GeneOntology No.
betaine-homocysteine S-methyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
betaine + L-homocysteine = dimethylglycine + L-methionine
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
methyl group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Cysteine and methionine metabolism
-
-
glycine betaine degradation I
-
-
glycine betaine degradation II (mammalian)
-
-
Glycine, serine and threonine metabolism
-
-
L-methionine salvage from L-homocysteine
-
-
Metabolic pathways
-
-
methionine metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
trimethylammonioacetate:L-homocysteine S-methyltransferase
-
CAS REGISTRY NUMBER
COMMENTARY hide
9029-78-1
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
Mus musculus C57BL/6J
C57BL/6J mice
-
-
Manually annotated by BRENDA team
-
Uniprot
Manually annotated by BRENDA team
Rattus norvegicus Sprague-Dawley
strain Sprague-Dawley
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
-
involved in homocystein 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
dimethylacetothetin + L-homocysteine
? + L-methionine
show the reaction diagram
-
-
-
?
glycinebetaine + L-homocysteine
? + L-methionine
show the reaction diagram
L-Asp + dimethylsulfonioacetate
?
show the reaction diagram
L-homocysteine + betaine
dimethylglycine + L-methionine
show the reaction diagram
L-homocysteine + betaine
L-methionine + dimethylglycine
show the reaction diagram
L-homocysteine + S-methyl-L-methionine
2 L-methionine
show the reaction diagram
-
-
-
-
?
L-homocysteine + S-methyl-L-methionine
?
show the reaction diagram
L-homocysteine + S-methylmethionine
2 L-methionine
show the reaction diagram
-
-
-
-
?
L-homocysteine + S-methylmethionine
?
show the reaction diagram
-
BHMT-2 uses S-methylmethionine as a methyl donor for the methylation of homocysteine. Unlike BHMT, BHMT-2 can not use betaine
-
-
?
N,N,N-trimethylglycine + L-homocysteine
N,N-dimethylglycine + L-methionine
show the reaction diagram
additional information
?
-
-
BHMT can utilize S-methylmethionine as a substrate in vitro with a very low affinity
-
-
-
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
L-homocysteine + betaine
L-methionine + dimethylglycine
show the reaction diagram
N,N,N-trimethylglycine + L-homocysteine
N,N-dimethylglycine + L-methionine
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
K+
-
Km value around 0.1 mM. The presence of potassium ions lowers the apparent KM of the enzyme for homocysteine, but it does not affect the apparent KM for betaine or the apparent kcat for either substrate
Zinc
wild-type enzyme contains zinc. Mutant enzymes H338A, R346A, W352A, R361A, P362A, Y363A, N364A, P365A maintain normal or near-normal ability to bind zinc
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(2S,11RS)-5-thia-2,11-diamino-8,8-dimethyldodecanedioic acid
-
more than 90% inhibition at 0.02 mM
(2S,11RS)-5-thia-2,11-diaminododecanedioic acid
-
about 60% inhibition at 0.02 mM
(2S,11S)-5,8-dithia-2,11-diaminododecanedioic acid
-
about 60% inhibition at 0.02 mM
(2S,5RS,8RS,11S)-5,8-dithia-2,11-diaminododecanedioic acid 5,8-dioxide
-
about 20% inhibition at 0.02 mM
(2S,8RS,11RS)-5-thia-2,11-diamino-8-methyldodecanedioic acid
-
competitive, more than 90% inhibition at 0.02 mM
(R,S)-2-(3-amino-3-carboxy-propylsulfanyl)-benzoic acid
-
ca. 43% inhibition at 0.02 mM
(R,S)-2-amino-4-(2-carboxy-ethyldisulfanyl)-butyric acid
-
ca. 30% inhibition at 0.02 mM
(R,S)-2-amino-4-(2-carboxymethylsulfanyl-ethylsulfanyl)-butyric acid
-
100% inhibition at 0.02 mM, very potent inhibitor and one of the strongest ever reported
(R,S)-2-amino-4-(2-carboxymethylsulfinyl-ethylsulfanyl)-butyric acid
-
ca. 90% inhibition at 0.02 mM
(R,S)-2-amino-4-(2-phosphonomethoxy-ethylsulfanyl)-butyrate
-
ca. 26% inhibition at 0.02 mM
(R,S)-2-amino-4-(3-carboxy-propylsulfanyl)-butyric acid
-
ca. 20% inhibition at 0.02 mM
(R,S)-2-amino-4-(4-carboxymethyl-benzylsulfanyl)-butyric acid
-
ca. 10% inhibition at 0.02 mM
(R,S)-2-amino-4-(4-phosphono-butylsulfanyl)-butyric acid
-
ca. 98% inhibition at 0.02 mM
(R,S)-2-amino-4-methylsulfanylmethylsulfanyl-butyric acid
-
ca. 21% inhibition at 0.02 mM
(R,S)-2-amino-4-[(phosphonomethyl-carbamoyl)-methylsulfanyl]-butyrate
-
ca. 9% inhibition at 0.02 mM
(R,S)-3-(3-amino-3-carboxy-propylsulfanyl)-benzoic acid
-
ca. 13% inhibition at 0.02 mM
(R,S)-4-(3-amino-3-carboxy-propylsulfanyl)-benzoic acid
-
ca. 99% inhibition at 0.02 mM
(R,S)-5-(3-amino-3-carboxy-propylsulfanyl)-pentanoic acid
-
100% inhibition at 0.02 mM, very potent inhibitor and one of the strongest ever reported, competitive inhibition with respect to betaine binding
(R,S)-5-(3-amino-3-carboxy-propylsulfinyl)-pentanoic acid
-
ca. 97% inhibition at 0.02 mM
(R,S)-5-(3-amino-3-carboxy-propylsulfonyl)-pentanoic acid
-
ca. 29% inhibition at 0.02 mM
(R,S)-5-(3-amino-3-carboxypropylsulfanyl)-pentanoic acid
-
(R,S)-6-(3-amino-3-carboxy-propylsulfanyl)-hexanoic acid
-
100% inhibition at 0.02 mM, very potent inhibitor and one of the strongest ever reported
(R,S,R,S)-2-amino-4-(2-amino-2-carboxy-ethylsulfinyl)-butyric acid
-
ca. 10% inhibition at 0.02 mM
(RS)-2-amino-4-[(2-carboxyethylthio)methylthio]butanoic acid
-
97.6% inhibition at 0.02 mM
(RS)-2-amino-4-[(3-carboxypropyl)disulfanyl]butanoic acid
-
19.1% inhibition at 0.02 mM
(RS)-2-amino-4-[2-(carboxymethylamino)ethylthio]butanoic acid
-
37.1% inhibition at 0.02 mM
(RS)-2-amino-4-[2-(R)-(1-carboxyethylamino)ethylthio]butanoic acid
-
15.5% inhibition at 0.02 mM
(RS)-2-amino-4-[2-(S)-(1-carboxyethylamino)ethylthio]butanoic acid
-
19.8% inhibition at 0.02 mM
(RS)-2-amino-4-[2-[(carboxymethyl)(methyl)amino]ethylthio]-butanoic acid
-
98.5% inhibition at 0.02 mM
(RS)-2-amino-4-[3-[(carboxymethyl)(methyl)amino]propylthio]butanoic acid
-
79.01% inhibition at 0.02 mM
(RS)-2-amino-5-(3-carboxypropylthio)pentanoic acid
-
5.4% inhibition at 0.02 mM
(RS)-2-aminodecanedioic acid
-
2.11% inhibition at 0.02 mM
(RS)-2-[[2-(3-amino-3-carboxypropylthio)ethyl]dimethylammonium]acetate
-
23.8% inhibition at 0.02 mM
(RS)-2-[[3-(3-amino-3-carboxypropylthio)propyl]dimethylammonio]acetate
-
3.3% inhibition at 0.02 mM
(RS)-5-(3-amino-3-carboxypropoxy)pentanoic acid
-
9.8% inhibition at 0.02 mM
(RS)-5-(3-amino-3-carboxypropylselanyl)pentanoic acid
-
complete inhibition at 0.02 mM
(RS)-5-(3-amino-3-carboxypropylthio)-3,3-dimethylpentanoic acid
-
highly potent inhibitor of BHMT, complete inhibition at 0.02 mM
(RS)-5-(3-amino-3-carboxypropylthio)-3-methylpentanoic acid
-
highly potent inhibitor of BHMT, complete inhibition at 0.02 mM
3,3-Dimethylbutyrate
-
competitive to betaine
5-(3-amino-3-carboxy-propylsulfanyl)-pentanoic acid
-
-
-
5-[(3-amino-3-carboxypropyl)sulfanyl]pentanoic acid
-
complete inhibition at 0.02 mM
5-[3-(R,S)-3-amino-3-(hydroxyphosphorylpropyl)sulfanyl]-3,3-dimethylpentanoic acid
-
-
-
5-[3-(R,S)-3-amino-3-(hydroxyphosphorylpropyl)sulfanyl]pentanoic acid
-
-
-
Ac-Val-Ala-Leu-His-NH2
-
0.1 mM, 25.9% inhibition
Ac-Val-DL-Ala-psi[(PO2-)-CH2]-DL-Leu-His-NH2
-
0.1 mM, 57.4% inhibition
Ac-Val-DL-Phe-psi[(PO2-)-CH2]-DL-Ala-His-NH2
-
0.1 mM, 8.7% inhibition
Ac-Val-DL-Phe-psi[(PO2-)-CH2]-DL-Leu-His-NH2
-
0.1 mM, 53.7% inhibition
actinomycin D
-
leads to a decay of Bhmt mRNA, irrespective of the ambient osmolarity, at 120 min after addition, Bhmt mRNA levels are significantly decreased under hyperosmotic conditions, compared with the Bhmt mRNA levels found under the respective normo- or hypoosmotic condition
AdoMet
Betaine aldehyde
-
100% inhibition at 2.5 mM
Butyrate
choline
-
60% inhibition at 5 mM
D-Cystine
-
-
dimethylglycine
dimethylsulfonioacetate
dimethylsulfoniopropionate
H2O2
-
causes a loss of catalytic Zn and a correlative loss of activity, irreversible
Isovalerate
-
competitive to betaine
L-Asp
-
10 mM Asp inhibits BHMT
L-cysteine
-
-
L-cystine
-
-
methionine
methyl methanethiosulfonate
-
causes a loss of catalytic Zn and a correlative loss of activity. Addition of beta-mercaptoethanol and exogenous Zn after methyl methanethiosulfonate treatment restores activity
N,N-dimethylglycine
-
70% inhibition at 5 mM
NaCl
-
100% inhibition above 200 mM
Pinanyl N,N,N-trimethylaminomethane boronate
-
substrate analogue
-
S-(delta-Carboxybutyl)-DL-homocysteine
S-(delta-carboxybutyl)-DL-homocysteine-sulfoxide
-
at 2 h after injection, there is a modest reduction in BHMT activity and a 90% increase in plasma total homocysteine
S-(delta-carboxybutyl)-L-homocysteine
S-adenosyl-L-ethionine
-
irreversible, S-adenosyl-L-homocysteine and L-homocysteine prevent, but not DL-homocysteine, GSH, DTT or L-cysteine
S-adenosyl-L-homocysteine
-
non-linear/competitive to homocysteine, mixed/non-competitive to betaine
S-adenosyl-L-methionine
Val-DL-Ala-psi[(PO2-)-CH2]-DL-Leu-His-NH2
-
0.1 mM, 70.6% inhibition
Val-DL-Phe-psi[(PO2-)-CH2]-DL-Leu-His-NH2
-
0.1 mM, 80.3% inhibition
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
beta-mercaptoethanol
-
using a redox-inert methyl acceptor, it is shown that BHMT requires a thiol reducing agent for activity. Short-term exposure of BHMT to reducing agent-free buffer inactivates the enzyme without causing any loss of its catalytic zinc. Activity can be completely restored by the re-addition of a thiol reducing agent
betaine
casein
-
included in liquid diets
choline
D-Cysteine
-
increase of product formation
dithiothreitol
-
-
ethanol
Insulin
-
is effective in counteracting the stimulation of BHMT activity and mRNA levels produced by triamcinolone in rat hepatoma H4IIE cells
-
L-cysteine
-
stimulation at low concentration
L-methionine
-
independent of dietary choline, supplemental methionine increases hepatic BHMT activity by ca. 30%
methionine
-
-
S-[(2R)-2-amino-2-carboxyethyl]-L-homocysteine
-
-
streptozotocin
-
increases BHMT activity and mRNA levels in the liver
Triamcinolone
-
-
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.002 - 2.4
betaine
0.155
dimethylacetothetin
-
0.008
DL-homocysteine
-
-
4.3
glycinebetaine
-
-
10
L-Asp
-
-
0.004 - 2.3
L-homocysteine
0.76 - 3.4
S-methyl-L-methionine
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000082 - 88
betaine
0.00023 - 0.0378
L-homocysteine
0.0044 - 38
S-methyl-L-methionine
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.386 - 0.716
betaine
0.108 - 2.52
L-homocysteine
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000077
(2S,8RS,11RS)-5-thia-2,11-diamino-8-methyldodecanedioic acid
-
IC50 for betaine-homocysteine S-methyltransferase about 0.077 mM, pH 7.5, 37C
0.000012
(R,S)-5-(3-amino-3-carboxy-propylsulfanyl)-pentanoic acid
-
-
0.045
Pinanyl N,N,N-trimethylaminomethane boronate
-
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000096
(R,S)-2-amino-4-(2-carboxymethylsulfanyl-ethylsulfanyl)-butyric acid
Homo sapiens
-
-
0.0057
(R,S)-2-amino-4-(4-phosphono-butylsulfanyl)-butyric acid
Homo sapiens
-
-
0.007
(R,S)-4-(3-amino-3-carboxy-propylsulfanyl)-benzoic acid
Homo sapiens
-
-
0.000087
(R,S)-5-(3-amino-3-carboxy-propylsulfanyl)-pentanoic acid
Homo sapiens
-
-
0.005
(R,S)-5-(3-amino-3-carboxy-propylsulfinyl)-pentanoic acid
Homo sapiens
-
-
0.0002
(R,S)-6-(3-amino-3-carboxy-propylsulfanyl)-hexanoic acid
Homo sapiens
-
-
0.00326
(RS)-2-amino-4-[(2-carboxyethylthio)methylthio]butanoic acid
Homo sapiens
-
in 50 mM potassium phosphate buffer, at pH 7.5, at 37C
0.0025
(RS)-2-amino-4-[2-[(carboxymethyl)(methyl)amino]ethylthio]-butanoic acid
Homo sapiens
-
in 50 mM potassium phosphate buffer, at pH 7.5, at 37C
0.000649
(RS)-5-(3-amino-3-carboxypropylselanyl)pentanoic acid
Homo sapiens
-
in 50 mM potassium phosphate buffer, at pH 7.5, at 37C
0.000084
(RS)-5-(3-amino-3-carboxypropylthio)-3,3-dimethylpentanoic acid
Homo sapiens
-
in 50 mM potassium phosphate buffer, at pH 7.5, at 37C
0.000139
(RS)-5-(3-amino-3-carboxypropylthio)-3-methylpentanoic acid
Homo sapiens
-
in 50 mM potassium phosphate buffer, at pH 7.5, at 37C
0.000093
5-(3-amino-3-carboxy-propylsulfanyl)-pentanoic acid
Homo sapiens
-
pH not specified in the publication, temperature not specified in the publication
-
0.0011
5-[3-(R,S)-3-amino-3-(hydroxyphosphorylpropyl)sulfanyl]-3,3-dimethylpentanoic acid
Homo sapiens
-
pH not specified in the publication, temperature not specified in the publication
-
0.005
5-[3-(R,S)-3-amino-3-(hydroxyphosphorylpropyl)sulfanyl]pentanoic acid
Homo sapiens
-
pH not specified in the publication, temperature not specified in the publication
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.000103
-
-
0.000557
-
-
0.00084
-
-
0.0025
-
kidney
0.0392
-
liver
0.314
-
-
2
-
mutant G27S, pH 7.5, 37C; mutant G27S, presence of 150 mM K+, pH 7.5, 37C
5
-
mutant D26A, pH 7.5, 37C
18
-
mutant D26A, presence of 150 mM K+, pH 7.5, 37C
100
-
wild-type, pH 7.5, 37C
940
-
wild-type, presence of 150 mM K+, pH 7.5, 37C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
-
assay at
additional information
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
10
-
pI-value of about 10
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
BHMT expression is observed in the amygdala of basal ganglia and the cerebral cortex where BHMT levels are clearly elevated during torpor
Manually annotated by BRENDA team
-
-
Manually annotated by BRENDA team
BHMT expression is observed in the amygdala of basal ganglia and the cerebral cortex where BHMT levels are clearly elevated during torpor
Manually annotated by BRENDA team
-
low BHMT activity is detected in the G30 fetus, and slightly increased levels of activity are observed in the liver from G45 and G90 fetuses
Manually annotated by BRENDA team
-
hyperosmolarity (405 mosmol/l) suppresses Bhmt mRNA expression whereas hypoosmotic (205 mosmol/l) conditions induce Bhmt mRNA expression. Like hyperosmotic NaCl, hyperosmotic raffinose but not hyperosmotic urea suppresses Bhmt mRNA expression. Osmosensitivity of Bhmt mRNA expression is impaired by inhibitors of tyrosine kinases and cyclic nucleotide-dependent kinases
Manually annotated by BRENDA team
-
-
Manually annotated by BRENDA team
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40300
-
sequence analysis of BHMT2
45000
-
about 45000 Da, SDS-PAGE
270000
350000
-
gel filtration
additional information
-
amino acid composition
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
mutant enzmyes R346A and W352A with negligible activity
hexamer
octamer
-
8 * 45000, SDS-PAGE
tetramer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
the enzyme can be modified by tissue-type transglutaminase and its activity is regulated repressively by the modification, especially by the cross-linking. This regulatory reaction might be involved in the regulation of homocysteine metabolism in the liver
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
by multiple anomalous diffraction
-
molecular dynamics simulations predict that K+ ions interact with residues Asp26 and/or Glu159. Crystal structure of BHMT bound to homocysteine confirms these sites of interaction and reveals further contacts between K+ ions and BHMT residues Gly27, Gln72, Gln247, and Gly298
-
recombinant enzyme
-
crystals with P2(1) symmetry, assymetric unit contains the whole functional tetramer showing point symmetry 222
-
recombinant enzyme
-
dominant structural feature of wild type BHMT is an (betaalpha)8 barrel. A modeled structure of truncated BHMT suggests that this protein would assume a horseshoe fold and lack methyltransferase activity
-
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5
-
above, 4 h stable at 37C
441234
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
significant heat stability
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
divalent cations and 2-mercaptoethanol do not stabilize during purification
-
glycerol does not stabilize
-
lyophilization inactivates
-
mechanism of unfolding by urea includes two intermediate states, a tetramer and a monomer. Dissociation is concomitant with alterations in the dimerization arm and the loop connecting the C-terminal helix, and intermediate conformations originate from perturbations in these structures
-
partial digestion of ligand-free enzyme with trypsin produces two large peptides, excising a seven residue peptide with loop L2. Carboxybutylhomocysteine but not methionine slows proteolysis by trypsin
-
substrates stabilize during purification
-
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
the enzyme is susceptible to conformation-dependent oxidative inactivation. When BHMT is incubated with up to 78 mM t-butyl hydroperoxide, no appreciable amount of ZN or activity is lost, indicating that oxidation of surface met residues are not involved in the oxidative inactivation of BHMT. Methyl methanethiosulfonate and H2O2, cause a loss of catalytic Zn and a correlative loss of activity. Addition of beta-mercaptoethanol and exogenous Zn after methyl methanethiosulfonate treatment restores activity, but oxidation due to H2O2 is irreversible. The L2 loop is involved in the conformational change associated with accupancy at the betaine binding site. This conformational change and/or occupancy at both ligand binding sites protects the enzyme from oxidative inactivation
-
657875
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 2 weeks in the presence of substrates
-
-20C, 40% loss of activity after 4 weeks in the presence of substrates
-
-20C, 70% loss of activity after 10 weeks in the presence of substrates
-
-20C, many months in 50% v/v glycerol
-
-20C, over a year in 50% v/v glycerol
-
stable for a minimum of one month at 4C in a mixture with human recombinant betaine-homocysteine S-methyltransferase (copurified with human recombinant betaine-homocysteine S-methyltransferase)
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
FPLC-chromatofocusing, 3 active forms
-
partial
-
stabilized by copurification with human recombinant betaine-homocysteine S-methyltransferase, chitin affinity chromatography
-
two forms of the enzyme
-
wild-type and mutant enzymes are overexpressed in Escherichia coli and purified using the ImpactTM T7 system
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
a 2.7 kb 5'-flanking region of the human BHMT gene cloned between MluI and SmaI sites of a promoterless pGL-3-basic vector creating the recombinant plasmid -2698/+33 BHMT-LUC, expression in HepG2 cells
expressed in COS-1 cells and in Escherichia coli; impossible to express BHMT2 in mammalian cells, protein aggregates after bacterial expression. BHMT2 is rapidly degraded in a rabbit reticulocyte lysate, but it can be stabilized by cotransfection of COS-1 cells with BHMT and, after cotransfection, it coprecipitates with BHMT
-
expressed in Escherichia coli BL21(DE3) (coexpression of human recombinant betaine-homocysteine S-methyltransferase)
-
expressed in Hep-G2 cell
-
glutathione S-transferase-BHMT fusion protein is expressed in HEK-293, T98G, A-10, MCF-7, H-1299, C2C12, and NIH-3T3 cells
-
transiently expressed in HepG2-cells and primary mouse hepatocytes
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
corticosteroids down-regulate in kidney expression and activity
-
corticosteroids stimulate activity in the liver and also up-regulate the gene expression
-
decreased expression after exogenous addition of all-trans-retinoic acid
-
during fetal development, a total of ten splice variants of BHMT are expressed at varying levels across a wide range of porcine tissues. Two variants contain an identical ORF that encodes a C-terminal truncated form of BHMT
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significant upregulation of betaine homocysteine methyltransferase-1 in livers of rats fed a betaine supplemented ethanol diet
-
the activity of hepatic BHMT significantly increases with increase in both dietary casein level and dietary methionine level, hepatic level of mRNA for BHMT is parallel to the enzyme activity
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A66V
-
nonsynonymous SNP identified using 240 DNA samples from four ethnic groups
Arg16Cys
-
nonsynonymous SNP identified using 240 DNA samples from four ethnic groups, Km (mM): 0.0139 (betaine), 0.0076 (L-homocysteine)
Arg239Gln
-
nonsynonymous SNP identified using 240 DNA samples from four ethnic groups, Km (mM): 0.012 (betaine), 0.0158 (L-homocysteine)
C104A
-
site-directed mutagenesis is used to investigate whether the loss of the DMSA-Asp activity of BHMT when in the absence of a reducing agent is due to the oxidation of an essential thiol within the protein. By individual mutation of each of the five Cys residues not involved in Zn binding to Ala, it is shown that the resulting mutants are as active as wild-type enzyme when in the presence of beta-mercaptoethanol with the DMSA-Asp assay
C131A
-
site-directed mutagenesis is used to investigate whether the loss of the DMSA-Asp activity of BHMT when in the absence of a reducing agent is due to the oxidation of an essential thiol within the protein. By individual mutation of each of the five Cys residues not involved in Zn binding to Ala, it is shown that the resulting mutants are as active as wild-type enzyme when in the presence of beta-mercaptoethanol with the DMSA-Asp assay
C186A
-
site-directed mutagenesis is used to investigate whether the loss of the DMSA-Asp activity of BHMT when in the absence of a reducing agent is due to the oxidation of an essential thiol within the protein. By individual mutation of each of the five Cys residues not involved in Zn binding to Ala, it is shown that the resulting mutants are as active as wild-type enzyme when in the presence of beta-mercaptoethanol with the DMSA-Asp assay
C201A
-
site-directed mutagenesis is used to investigate whether the loss of the DMSA-Asp activity of BHMT when in the absence of a reducing agent is due to the oxidation of an essential thiol within the protein. By individual mutation of each of the five Cys residues not involved in Zn binding to Ala, it is shown that the resulting mutants are as active as wild-type enzyme when in the presence of beta-mercaptoethanol with the DMSA-Asp assay
C217A
-
the mutation reduces zinc binding by 95% while abrogating catalytic activity, the mutation has no effect on the fold increase of GST-BHMT proteolytic fragment in the absence of nutrients
C256A
-
site-directed mutagenesis is used to investigate whether the loss of the DMSA-Asp activity of BHMT when in the absence of a reducing agent is due to the oxidation of an essential thiol within the protein. By individual mutation of each of the five Cys residues not involved in Zn binding to Ala, it is shown that the resulting mutants are as active as wild-type enzyme when in the presence of beta-mercaptoethanol with the DMSA-Asp assay
Cys217Ala
-
complete loss of activity, reduction in zinc binding, identification of zinc binding motif
Cys299Ala
-
complete loss of activity, reduction in zinc binding, identification of zinc binding motif
Cys300Ala
-
complete loss of activity, reduction in zinc binding, identification of zinc binding motif
D26A
-
almost 3fold decrease in stimulation by K+
DELTA325-406
truncation mutatnt does not express well in Escherichia coli and is inactive
DELTA371-406
truncation mutant does not express well in Escherichia coli and is inactive
E159A
-
complete loss of activity
E159Q
-
complete loss of activity
E266A
near-normal catalytic activity
G199S
-
in vascular patients with hyperhomocysteinemia
G27S
-
no stimulation by K+
G28S
-
very small decrease in stimulation by K+
G742A
-
in an ongoing, multicenter, case-control study including women with a clinical diagnosis of abruption an association between the homozygous mutant form of BHMT (742G to A) polymorphism and an increased risk for placental abruption is shown
Gly199Ser
-
nonsynonymous SNP identified using 240 DNA samples from four ethnic groups, Km (mM): 0.0139 (betaine), 0.0206 (L-homocysteine)
H338A
normal or near-normal ability to bind zinc, 10% of the wild-type activity
N364A
normal or near-normal ability to bind zinc, near-normal catalytic activity
P362A
normal or near-normal ability to bind zinc, near-normal catalytic activity
P365A
normal or near-normal ability to bind zinc, near-normal catalytic activity
Pro197Ser
-
nonsynonymous SNP identified using 240 DNA samples from four ethnic groups, Km (mM): 0.0213 (betaine), 0.0216 (L-homocysteine)
R239Q
-
no significant association with the severity and extent of hyperhomocysteinemia
R346A
normal or near-normal ability to bind zinc, negligible activity, elution as dimer, aberrant crosslinking properties
R361A
normal or near-normal ability to bind zinc, near-normal catalytic activity
T218M
-
nonsynonymous SNP identified using 240 DNA samples from four ethnic groups
V155F
-
nonsynonymous SNP identified using 240 DNA samples from four ethnic groups
V237M
-
nonsynonymous SNP identified using 240 DNA samples from four ethnic groups
Y363A
normal or near-normal ability to bind zinc, near-normal catalytic activity
A119G
-
1.9fold reduced turnover-number for betaine and L-homocysteine, 1.6fold reduced KM-value for L-homocysteine, 1.1fold decrease in Km-value for betaine
C186A
-
1.8fold reduced turnover-number for betaine and L-homocysteine, 4.3fold reduced KM-value for L-homocysteine, 1.8fold increase in Km-value for betaine
C186S
-
2.3fold reduced turnover-number for betaine and L-homocysteine, 1.4fold reduced KM-value for L-homocysteine, 111.6fold decrease in Km-value for betaine
D26A
-
2.7fold reduced turnover-number for betaine and L-homocysteine, 1.7fold reduced KM-value for L-homocysteine, 3.5fold increase in Km-value for betaine
D26I
-
67fold reduced turnover-number for betaine and L-homocysteine, maximal velocity is 6.5fold lower than the wild-type value
E159G
-
60fold reduced turnover-number for betaine and L-homocysteine, maximal velocity is 61.2fold lower than the wild-type value
E159K
-
170fold reduced turnover-number for betaine and L-homocysteine, maximal velocity is 169.7fold lower than the wild-type value
E21A
-
2.3fold reduced turnover-number for betaine and L-homocysteine, 6fold reduced KM-value for L-homocysteine, 2.4fold reduced Km-value for betaine
E21K
-
1.6fold reduced turnover-number for betaine and L-homocysteine, 1.4fold reduced KM-value for L-homocysteine, 1.7fold reduced Km-value for betaine
F74A
-
1.9fold reduced turnover-number for betaine and L-homocysteine, 1.9fold reduced KM-value for L-homocysteine, 2.9fold increase in Km-value for betaine
R346A
-
shows alterations in the association state
T184G
-
2.4fold reduced turnover-number for betaine and L-homocysteine, 3.19fold reduced KM-value for L-homocysteine, 1.6fold increase in Km-value for betaine
T73G
-
3.1fold reduced turnover-number for betaine and L-homocysteine, 2.2fold reduced KM-value for L-homocysteine, 1.3fold reduced Km-value for betaine
W352A
-
shows alterations in the association state
Y363A
-
shows alterations in the association state
Y77A
-
14fold reduced turnover-number for betaine and L-homocysteine, maximal velocity is 13.7fold lower than the wild-type value
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
-
synthesis of a series of S-substituted derivatives of homocysteine as potential inhibitors of human recombinant BHMT, some of these compounds are very potent inhibitors, having IC50 values in the nanomolar range
medicine
molecular biology
additional information