Information on EC 2.1.2.10 - aminomethyltransferase

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

EC NUMBER
COMMENTARY hide
2.1.2.10
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RECOMMENDED NAME
GeneOntology No.
aminomethyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
[protein]-S8-aminomethyldihydrolipoyllysine + tetrahydrofolate = [protein]-dihydrolipoyllysine + 5,10-methylenetetrahydrofolate + NH3
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C-N bond cleavage
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-
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methyl group transfer
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-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
glycine cleavage
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glycine metabolism
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Glycine, serine and threonine metabolism
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Metabolic pathways
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NIL
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One carbon pool by folate
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SYSTEMATIC NAME
IUBMB Comments
[protein]-S8-aminomethyldihydrolipoyllysine:tetrahydrofolate aminomethyltransferase (ammonia-forming)
A component, with EC 1.4.4.2 glycine dehydrogenase (decarboxylating) and EC 1.8.1.4, dihydrolipoyl dehydrogenanse, of the glycine cleavage system, formerly known as glycine synthase. The glycine cleavage system is composed of four components that only loosely associate: the P protein (EC 1.4.4.2), the T protein (EC 2.1.2.10), the L protein (EC 1.8.1.4) and the lipoyl-bearing H protein [3].
CAS REGISTRY NUMBER
COMMENTARY hide
37257-08-2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
male Wistar rats
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
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aminomethyltransferase is a component of the T-protein, which is part of a multienzyme system composed of four proteins termed P-, H-, T-, and L-protein. T-protein/aminomethyltransferase degrades the aminomethyl moiety to ammonia and 5,10-methylentetrahydrofolate in the presence of tetrahydrofolate, leaving dihydrolipoate-bearing H-protein
physiological function
additional information
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T-protein in complex with dihydrolipoate-bearing H-protein and 5-methyltetrahydrofolate, a complex mimicking the ternary complex in the reverse reaction, shows a highly interacting intermolecular interface limited to a small area and the protein-bound dihydrolipoyllysine arm inserted into the active site cavity of the T-protein. Arg292 of the T-protein is essential for complex assembly
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
reduced H-protein + 5,10-methylene-tetrahydrofolate + NH4Cl
?
show the reaction diagram
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-
-
-
?
reduced H-protein + 5,10-methylenetetrahydropteroyltetraglutamate + NH4Cl
?
show the reaction diagram
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-
-
-
?
S-aminomethyldihydrolipoylprotein + (6S)-tetrahydrofolate
dihydrolipoylprotein + (6R)-5,10-methylenetetrahydrofolate + NH3
show the reaction diagram
S-aminomethyldihydrolipoylprotein + tetrahydropteroyltetraglutamate
dihydrolipoylprotein + 5,10-methylenetetrahydropteroyltetraglutamate + NH3
show the reaction diagram
-
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better substrate than 5,10-methylenetetrahydrofolate, 6.5fold higher affinity for 5,10-methylenetetrahydropteroyltetraglutamate than for 5,10-methylenetetrahydrofolate
?
[protein]-S8-aminomethyldihydrolipoyllysine + tetrahydrofolate
[protein]-dihydrolipoyllysine + 5,10-methylenetetrahydrofolate + NH3
show the reaction diagram
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-
-
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r
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
S-aminomethyldihydrolipoylprotein + (6S)-tetrahydrofolate
dihydrolipoylprotein + (6R)-5,10-methylenetetrahydrofolate + NH3
show the reaction diagram
[protein]-S8-aminomethyldihydrolipoyllysine + tetrahydrofolate
[protein]-dihydrolipoyllysine + 5,10-methylenetetrahydrofolate + NH3
show the reaction diagram
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r
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
pyridoxal 5'-phosphate
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tetrahydrofolate
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
iodoacetamide
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5 mM, 70% inhibition
N-ethylmaleimide
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0.1 mM, 70% inhibition
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0677 - 0.151
5,10-methylenetetrahydrofolate
0.0092 - 0.0382
5,10-methylenetetrahydropteroyltetraglutamate
65.4 - 256
NH4Cl
0.00069 - 0.0169
reduced H-protein
0.0037
S-aminomethyldihydrolipoylprotein
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0.17
tetrahydrofolate
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-
additional information
additional information
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values for mutant enzymes
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.1 - 18.4
5,10-methylenetetrahydrofolate
8.75 - 11.6
5,10-methylenetetrahydropteroyltetraglutamate
3.9 - 20.9
NH4Cl
2.9 - 19.4
reduced H-protein
additional information
additional information
Escherichia coli
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values for mutant enzymes
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
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assay at
additional information
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pI: 9.8
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Bacillus subtilis (strain 168)
Bartonella henselae (strain ATCC 49882 / DSM 28221 / Houston 1)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
33000
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gel filtration
37000 - 38000
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sedimentation equilibrium centrifugation, gel filtration
45000
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gel filtration
57000
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T-protein in complex with H-protein, gel filtration
75000
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dynamic light scattering
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 30000, SDS-PAGE
monomer
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
aminomethyltransferase/T-protein in complex with dihydrolipoate-bearing H-protein and 5-methyltetrahydrofolate, a complex mimicking the ternary complex in the reverse reaction, purified recombinant wild-type and mutant T-proteins, hanging drop vapor diffusion method, mixing of 22-25 mg/ml proteins, T-protein and H protein, with an equal volume of reservoir solution containing 0.095 M 2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid, pH 7.5, 0.19 M calcium chloride, 26.6% v/v PEG 400, and 5% v/v glycerol, at 15C, 4-5 days to 2 weeks, soaking of the native crystals in mother liquid containing 1 mM (6S)-5-methyltetrahydrofolate at 15C for 4 h, X-ray diffraction structure determination and analysis at 2.0 A resolution, molecular replacement
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apoform and bound to 5-methyltetrahydrofolate
apoform, tetrahydrofolate complex, folinic acid complex and lipoic acid complex of enzyme, homology model of human enzyme
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
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1 min, 20% loss of activity
70
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1 min, complete loss of activity
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
lability of enzyme under the conditions of its purifications and storage
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 20 mM Tris-HCl buffer, pH 8.0, 10 mM 2-mercaptoethanol, 50% v/v glycerol, at least 2 months, stable
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-20C, 24 h, 60% loss of activity
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0-4C, 1 week, stable
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
641fold purification
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8.8fold purification
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purification of recombinant enzyme, expressed in Escherichia coli BL21 (DE3)pLysS, and of mutants
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Amt gene is located on chromosome 3p21 and contains 9 exons spanning about 6 kb of genomic sequence
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C-terminally His6-tagged T-protein wild-type and mutant enzymes expression in Escherichia coli strain BL21(DE3)pLysS
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cloning and sequencing of the Amt gene with 9 closely spaced exons that are contained within about 5 kb of genomic DNA, encoding a protein of 403 amino acids, cis-acting promoter is likely to be very short, Amt gene may be localized on chromosome 9F
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transfection to Leishmania infantum and Leishmania major
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
decrease of expression of isogene 1 of T-protein component of glcycine cleavage complex in amastigote growth
increased expression upon exposure to light
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response to heat stress in higher expression of isogene 1 of T-protein component of glcycine cleavage complex, inducible expression upon excess glycine
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D96N
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site-directed mutagenesis, both the glycine cleavage and synthesis activities are reduced to 34% compared to the wild-type enzyme
D96N/Y188F
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site-directed mutagenesis, the mutations abolish both the glycine cleavage and synthesis activities
D97N
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site-directed mutagenesis, the mutation abolishes both the glycine cleavage and synthesis activities
D97N/Y188F
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site-directed mutagenesis, the mutations abolish both the glycine cleavage and synthesis activities
K352E
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mutant with 2fold increased Km-values for folate substrates
K352Q
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mutant with 2fold increased Km-values for folate substrates
K352R
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no effect on Km-values
K75E
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mutant with 2.5fold increased Km-value for 5,10-methylenetetrahydrofolate and 8fold increased Km-value for 5,10-methylenetetrahydropteroyltetraglutamate
K78E
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mutant with 1.4fold increased Km-values for folate substrates
K81E
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mutant with 3fold increased Km-value for 5,10-methylenetetrahydrofolate and 16fold increased Km-value for 5,10-methylenetetrahydropteroyltetraglutamate
L6A
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in contrast to wild-type, quite susceptible to trypsinolysis, 4fold increase in Km-value for reduced H-protein
N113A
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site-directed mutagenesis, the mutation abolishes both the glycine cleavage and synthesis activities
N113A/R223A
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site-directed mutagenesis, the mutations abolish both the glycine cleavage and synthesis activities
N113D
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site-directed mutagenesis, the mutation abolishes both the glycine cleavage and synthesis activities
R223A
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site-directed mutagenesis, the mutation abolishes both the glycine cleavage and synthesis activities
R223K
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site-directed mutagenesis, the mutation abolishes both the glycine cleavage and synthesis activities
T4A
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2fold increase in Km-value for reduced H-protein
Y188F
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site-directed mutagenesis, both the glycine cleavage and synthesis activities are reduced to 83% compared to the wild-type enzyme
D276H
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nonketotic hyperglycinemia, rare mutation
E211K
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polymorphism occurring in patients with glycine encephalopathy, NKH, method for PCR-restriction enzyme analysis
G269D
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nonketotic hyperglycinemia, rare mutation
G47R
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nonketotic hyperglycinemia, rare mutation
H42R
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present in many nonketotic hyperglycinemia affected members of an extended Israeli-Arab kindred
N117I
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mutant may cause nonketotic hygerglycinemia
N145I
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nonketotic hyperglycinemia, substitution of conserved N, patient has servere neonatal presentation and died in the newborn period
Q192X
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nonketotic hyperglycinemia, premature stop codon
R296H
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mutation occurring in patients with glycine encephalopathy, NKH, method for PCR-restriction enzyme analysis
R318R
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polymorphism occurring in patients with glycine encephalopathy, NKH, method for PCR-restriction enzyme analysis
R320H
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allele frequency of 7% for R320H of T-protein in 50 patients with enzymatic confirmation of their diagnostics of nonketotic hyperglycinemia
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine
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