Information on EC 4.3.1.2 - methylaspartate ammonia-lyase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
4.3.1.2
-
RECOMMENDED NAME
GeneOntology No.
methylaspartate ammonia-lyase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
L-threo-3-methylaspartate = mesaconate + NH3
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
elimination
elimination of NH3
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
C5-Branched dibasic acid metabolism
-
-
Glyoxylate and dicarboxylate metabolism
-
-
L-glutamate degradation VI (to pyruvate)
-
-
Metabolic pathways
-
-
methylaspartate cycle
-
-
glutamate and glutamine metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
L-threo-3-methylaspartate ammonia-lyase (mesaconate-forming)
A cobalamin protein.
CAS REGISTRY NUMBER
COMMENTARY hide
9033-26-5
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Bacterium cadaveris
-
-
-
Manually annotated by BRENDA team
TPU 0011
-
-
Manually annotated by BRENDA team
strain YG-0504
-
-
Manually annotated by BRENDA team
JCM 1658; JCM 1659
-
-
Manually annotated by BRENDA team
JCM 1658
-
-
Manually annotated by BRENDA team
JCM 1659
-
-
Manually annotated by BRENDA team
NCIMB 10466
-
-
Manually annotated by BRENDA team
NCIMB 232
-
-
Manually annotated by BRENDA team
NCIMB 9525
-
-
Manually annotated by BRENDA team
IAM 1133
-
-
Manually annotated by BRENDA team
IAM 1133
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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
(2E)-2-butylbut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 0%, mutant L384A: conversion 59%
-
-
r
(2E)-2-ethylbut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 74%, mutant L384A: conversion 76%
-
-
r
(2E)-2-hexylbut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 0%, mutant L384A: conversion 53%
-
-
r
(2E)-2-methylbut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 80%, mutant L384A: conversion 75%
-
-
r
(2E)-2-pentylbut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 0%, mutant L384A: conversion 52%
-
-
r
(2E)-2-phenylbut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 0%, mutant L384A: conversion 90%
-
-
r
(2E)-2-propylbut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 57%, mutant L384A: conversion 67%
-
-
r
(2R,3S)-3-chloroaspartic acid
2-chloro-2-butenedioic acid
show the reaction diagram
(2S)-2-hydrazinosuccinic acid
fumarate + hydrazine
show the reaction diagram
-
-
-
-
(2S)-aspartate
fumarate + NH3
show the reaction diagram
-
-
-
?
(2S)-N,N-dimethylaspartic acid
fumarate + dimethylamine
show the reaction diagram
-
-
-
-
(2S)-N-ethylaspartic acid
fumarate + ethylamine
show the reaction diagram
-
-
-
-
(2S)-N-hydroxyaspartic acid
fumarate + hydroxylamine
show the reaction diagram
-
-
-
-
(2S)-N-methoxyaspartic acid
fumarate + methoxylamine
show the reaction diagram
-
-
-
-
(2S)-N-methylaspartic acid
fumarate + methylamine
show the reaction diagram
-
-
-
-
(2S,3R)-3-ethylaspartic acid
?
show the reaction diagram
(2S,3S)-3-ethylaspartate
?
show the reaction diagram
-
-
-
?
(2S,3S)-3-ethylaspartic acid
?
show the reaction diagram
(2S,3S)-3-methylaspartate
mesaconate + NH3
show the reaction diagram
(2S,3S)-N-hydroxy-3-ethylaspartic acid
ethylfumarate + hydroxylamine
show the reaction diagram
-
-
-
-
(2S,3S)-N-hydroxy-3-methylaspartic acid
mesaconate + hydroxylamine
show the reaction diagram
-
-
-
-
(2S,3S)-N-methoxy-3-methylaspartic acid
mesaconate + methoxylamine
show the reaction diagram
-
-
-
-
(2Z)-2-(benzyloxy)but-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 0%, mutant L384A: conversion 60%
-
-
r
(2Z)-2-(benzylsulfanyl)but-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 0%, mutant L384A: conversion 42%
-
-
r
(2Z)-2-(ethylsulfanyl)but-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 36%, mutant L384A: conversion 50%
-
-
r
(2Z)-2-(phenylsulfanyl)but-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 0%, mutant L384A: conversion 34%
-
-
r
(2Z)-2-ethoxybut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 46%, mutant L384A: conversion 43%
-
-
r
(2Z)-2-phenoxybut-2-enedioate + NH3
?
show the reaction diagram
-
wild-type: conversion 0%, mutant L384A: conversion 44%
-
-
r
(S)-aspartate
fumarate + NH3
show the reaction diagram
-
-
-
?
2-hydrazino-3-chlorosuccinic acid
chlorofumarate + hydrazine
show the reaction diagram
-
-
-
-
2-hydrazino-3-ethylsuccinic acid
ethylfumarate + hydrazine
show the reaction diagram
-
-
-
-
2-hydrazino-3-isopropylsuccinic acid
isopropylfumarate + hydrazine
show the reaction diagram
-
-
-
-
2-hydrazino-3-propylsuccinic acid
propylfumarate + hydrazine
show the reaction diagram
-
-
-
-
3-bromoaspartate
bromofumarate + NH3
show the reaction diagram
3-chloroaspartate
chlorofumarate + NH3
show the reaction diagram
3-fluoroaspartate
fluorofumarate + NH3
show the reaction diagram
-
-
not a good substrate
-
? + NH4Cl
?
show the reaction diagram
beta-ethylaspartate
?
show the reaction diagram
beta-isopropylaspartate
?
show the reaction diagram
-
-
-
-
-
beta-propylaspartate
?
show the reaction diagram
erythro-3-methylaspartate
?
show the reaction diagram
ethylfumarate + NH3
?
show the reaction diagram
hydrazino-3-methylsuccinic acid
mesaconate + hydrazine
show the reaction diagram
-
-
-
-
iso-propylfumarate + NH3
?
show the reaction diagram
-
-
-
-
-
L-aspartate
fumarate + NH3
show the reaction diagram
L-erythro-3-methylaspartate
mesaconate + NH3
show the reaction diagram
L-threo-3-methylaspartate
?
show the reaction diagram
L-threo-3-methylaspartate
mesaconate + NH3
show the reaction diagram
L-threo-3-methylaspartic acid
mesaconate + NH3
show the reaction diagram
assay at pH 9.0, 30C
-
-
?
mesaconate + (aminooxy)ethane
?
show the reaction diagram
-
mutant Q73A, conversion: 100%, yield: 61%
-
-
r
mesaconate + (aminooxy)methane
?
show the reaction diagram
-
mutant Q73A, conversion: 99%, excess of threo isomer over erythro isomer: more than 95%
-
-
r
mesaconate + 1-cyclopropylmethanamine
?
show the reaction diagram
mesaconate + 2-aminoethanol
?
show the reaction diagram
-
mutant Q73A, conversion: 37%, excess of threo isomer over erythro isomer: more than 95%
-
-
r
mesaconate + 2-methoxyethanamine
?
show the reaction diagram
mesaconate + 3-aminopropan-1-ol
?
show the reaction diagram
mesaconate + aminobutane
?
show the reaction diagram
mesaconate + aniline
?
show the reaction diagram
-
mutant Q73A, conversion: 6%, excess of threo isomer over erythro isomer: more than 95%
-
-
r
mesaconate + cyclobutanamine
?
show the reaction diagram
mesaconate + cyclohexanamine
?
show the reaction diagram
-
mutant Q73A, conversion: 21%, excess of threo isomer over erythro isomer: more than 95%
-
-
r
mesaconate + cyclopentanamine
?
show the reaction diagram
-
mutant Q73A, conversion: 36%, excess of threo isomer over erythro isomer: more than 95%
-
-
r
mesaconate + cyclopropanamine
?
show the reaction diagram
mesaconate + ethanamine
?
show the reaction diagram
mesaconate + ethane-1,2-diamine
?
show the reaction diagram
mesaconate + ethylamine
?
show the reaction diagram
-
-
-
?
mesaconate + hexanamine
?
show the reaction diagram
-
mutant Q73A, conversion: 17%, excess of threo isomer over erythro isomer: more than 95%
-
-
r
mesaconate + hydrazine
?
show the reaction diagram
-
-
-
?
mesaconate + hydroxylamine
?
show the reaction diagram
-
-
-
?
mesaconate + methanamine
?
show the reaction diagram
mesaconate + methoxylamine
?
show the reaction diagram
-
-
-
?
mesaconate + methylamine
?
show the reaction diagram
-
-
-
?
mesaconate + N-methylethane-1,2-diamine
?
show the reaction diagram
-
mutant Q73A, conversion: 80%, excess of threo isomer over erythro isomer: more than 95%
-
-
r
mesaconate + NH3
?
show the reaction diagram
-
-
-
?
mesaconate + NH3
L-threo-3-methylaspartate
show the reaction diagram
mesaconate + NH3
L-threo-3-methylaspartate + L-erythro-3-methylaspartate
show the reaction diagram
-
conversion 2h: 73% (wild-type), diastereomeric product ratio (threo:erythro) after 2h: 89:11
-
-
r
mesaconate + NH4Cl
methylaspartate + HCl
show the reaction diagram
-
-
-
-
r
mesaconate + pentanamine
?
show the reaction diagram
mesaconate + propan-2-amine
?
show the reaction diagram
-
mutant Q73A, conversion: 8%, excess of threo isomer over erythro isomer: more than 95%, enantiomeric excess: more than 99%
-
-
r
mesaconate + propanamine
?
show the reaction diagram
mesaconate + propane-1,3-diamine
?
show the reaction diagram
mesaconic acid + NH3
L-threo-3-methylaspartate
show the reaction diagram
assay at pH 9.0, 30C
-
-
r
methylfumarate + NH3
?
show the reaction diagram
-
-
-
?
N-3-dimethylaspartic acid
methylfumarate + methylamine
show the reaction diagram
-
-
-
-
N-methyl-3-ethylaspartic acid
ethylfumarate + methylamine
show the reaction diagram
-
-
-
-
NH3 + (2E)-2-methylbut-2-enedioate
(2S,3R)-3-methylaspartate
show the reaction diagram
-
-
minor product
r
NH3 + (2E)-2-methylbut-2-enedioate
(2S,3S)-3-methylaspartate
show the reaction diagram
-
-
major product
r
propylfumarate + NH3
?
show the reaction diagram
-
-
-
?
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
L-erythro-3-methylaspartate
mesaconate + NH3
show the reaction diagram
-
i.e. (2S,3R)-3-methylaspartate, slow syn elimination
-
r
L-threo-3-methylaspartate
?
show the reaction diagram
L-threo-3-methylaspartate
mesaconate + NH3
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Cd2+
-
activates
Cu2+
-
approx. 10% of Mg2+ activation
NH4+
-
required
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2,2'-dipyridyl
-
70% loss of activity in the deamination reaction of L-threo-3-methylaspartate
3-bromoaspartate
-
irreversible
4-nitrophenylhydrazine
-
90% inhibition after 200 min
5,5'-dithiobis(2-nitrobenzoate)
8-hydroxyquinoline
Al3+
-
5 mM AlCl3, 95% loss of activity
hydroxylamine
-
-
iodoacetate
-
1 mM, 86% loss of activity
methylhydrazine
-
-
N-ethylmaleimide
o-phenanthroline
-
24% loss of activity in the deamination reaction of L-threo-3-methylaspartate
p-chloromercuribenzoate
p-mercuribenzoate
-
-
Pb2+
-
5 mM PbCl2, 88% loss of activity
phenylhydrazine
Sr2+
-
competitive inhibition of Mg2+ activation of the enzyme
Zn2+
-
30% loss of activity in the deamination reaction of L-threo-3-methylaspartate
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Dimethylsulfoxide
NH4+
-
approx. 65% of K+ activation
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
7.9
(2R,3S)-3-chloroaspartic acid
-
-
5.2 - 40
(2S,3R)-3-methylaspartate
4.6 - 16
(2S,3S)-3-ethylaspartic acid
0.76
(2S,3S)-3-methylaspartate
-
-
0.51
(2S,3S)-methylaspartate
-
25C, pH 9.7
1.5
(S)-aspartate
-
25C, pH 9.7
0.58
L-threo-3-methylaspartate
-
recombinant 3-methylaspartase
1 - 1.8
L-threo-3-methylaspartic acid
0.2 - 60
mesaconate
0.13 - 14
mesaconic acid
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
16 - 78
(2S,3S)-3-ethylaspartic acid
240 - 467
L-threo-3-methylaspartate
0.01 - 89
L-threo-3-methylaspartic acid
0.6 - 61
mesaconate
0.25 - 61
mesaconic acid
additional information
additional information
Clostridium tetanomorphum
-
-
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4.9 - 35
(2S,3S)-3-ethylaspartic acid
95416
0.01 - 89
L-threo-3-methylaspartic acid
12085
0.04 - 87
mesaconate
3214
0.15 - 87
mesaconic acid
7232
additional information
L-threo-3-methylaspartic acid
12085
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
282
-
incubation with 5 mM EDTA, pH 7.0, 4C, for 3 h
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8 - 8.5
-
amination
8
-
maximal activity in TRIS/HCl buffer in the amination reaction of mesaconate
9.5
-
amination
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 11
-
pH 6.0: 10% of maximal activity, pH 11.0: 40% of maximal activity in the amination reaction of mesaconate
7 - 11
8.5 - 11
-
55% of maximal amination activity at pH 8.5, 75% of maximal amination activity at pH 11.0, 30% of maximal deamination activity at pH 8.5, 15% of maximal deamination activity at pH 11.0
additional information
-
-
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45
-
maximal activity in the deamination reaction of L-threo-3-methylaspartate
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45000
-
2 * 45000, SDS-PAGE
45538
-
2 * 45538, mass spectrometry
45539
-
2 * 45539, calculation from sequence of amino acid
46000
-
2 * 46000, SDS-PAGE
70000
-
gel filtration
79000
-
gel filtration
84000
-
gel filtration
88000
-
gel filtration
100000
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapour diffusion, well solution consists of 100 mM potassium phosphate, pH 8.0, and 32% ammonium sulfate, crystals diffract to 1.3 A
-
crystallized from ammonium sulfate
-
once recrystallized
-
sitting-drop vapour-diffusion, 0.0045 ml protein sample and 0.0045 ml precipitant, crystals are obtained using 20-25% polyethylene glycol 6000, 100 mM sodium acetate, pH 7.0, 25 mM Tris-HCl as precipitant solution and 16-22% ethylene glycol as additive, crystals diffract to 2.0 A
-
sitting-drop vapur diffusion against a well solution of 20-25% polyethylene glycol 6000, 100 mM sodium acetate, pH 7.0, 25 mM Tris-HCl pH 7.0 as precipitating solution and 16-22% ethylene glycol as additive, crystals diffract to 1.9 A
-
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 10
-
stable between, pH 5.0: inactivated rapidly below
6060
5.5 - 11
-
30C, 10 min, deamination reaction of L-threo-3-methylaspartate, 14% loss of activity
6059
6 - 8.5
-
stable between, pH 5.0: inactivated rapidly below
6060
additional information
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
-
0.1 mM potassium phosphate buffer, pH 7.0, 10 min, in the deamination reaction of (2S,3S)-3-methxylaspartate, 96.5% loss of activity
additional information
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
enzyme production is suppressed by an incorporation of aerobic cultivation before growth under oxygen-limited conditions
additional information
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
0-3C, in 0.5 M tetramethylammonium chloride, pH 6.8 for many months, with little loss of activity
-
4C, enzyme crystals retain full activity after 1 year
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
from whole cell lysates for protein identification using 2D-electrophoresis and LC-MS/MS
-
heat, ammonium sulfate, gel filtration, DEAE-Sepharose, ethanol
-
JCM 1658, JCM 1659
-
native and recombinant 3-methylaspartase
-
recombinant 3-methylaspartase
using metal affinity chromatography
YG-0504, TPU 0011, IFO 13539, IFO 13545, IFO 13546
-
YG-0601, IFO 3168, IFO 3848, NCIMB 232, NCIMB 9525, NCIMB 10466, NCTC 1707, NCTC 2815, NCTC 2818, NCTC 5845, NCTC 7381, NCTC10011, NCTC 10375, NCTC11354, NCTC 12028
-
YG-1002, IFO 13547
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli
-
expression in Escherichia coli
in Escherichia coli
-
overexpressed in Escherichia coli
overexpression in Escherichia coli
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C361A
-
kcat (mesaconate) decreased compared to wild-type, Km (mesaconate) increased compared to wild-type. Conversion 2h: 47% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 95:5 (89:11 wild-type)
C361K
-
mutant shows no reactivity. Conversion 2h: 10% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 95:5 (89:11 wild-type)
F170A
-
kcat (mesaconate) decreased compared to wild-type, Km (mesaconate) increased compared to wild-type. Conversion 2h: 63% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 95:5 (89:11 wild-type)
H194A
-
enhanced diastereoselectivities
K331A
-
enhanced diastereoselectivities
Q172A
-
kcat (mesaconate) decreased compared to wild-type, Km (mesaconate) increased compared to wild-type. Conversion 2h: 16% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 95:5 (89:11 wild-type)
Q172N
-
mutant shows no reactivity. Conversion 2h: 6% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 95:5 (89:11 wild-type)
Q329A
-
enhanced diastereoselectivities
Q73N
-
mutant shows very low-level amination activity preventing the measurement of kinetic parameters. Conversion 2h: 18% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 95:5 (89:11 wild-type)
T360A
-
kcat (mesaconate) decreased compared to wild-type, Km (mesaconate) increased compared to wild-type. Conversion 2h: 38% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 93:7 (89:11 wild-type)
T360S
-
kcat (mesaconate) decreased compared to wild-type, Km (mesaconate) increased compared to wild-type. Conversion 2h: 76% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 86:14 (89:11 wild-type)
Y356A
-
kcat (mesaconate) decreased compared to wild-type, Km (mesaconate) increased compared to wild-type. Conversion 2h: 24% (73% wild-type), diastereomeric product ratio (threo:erythro) after 2h: 95:5 (89:11 wild-type)
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
biotechnology
-
Fusobacterium varium can simultaneously utilize both glucose and L-glutamate as energy sources, intracellular concentrations of methylaspartate ammonia-lyase is elevated when the bacterium is cultured in media supplemented with excess L-glutamate
synthesis
-
enzyme is used for L-aspartic acid production
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