Information on EC 4.1.2.48 - low-specificity L-threonine aldolase

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

EC NUMBER
COMMENTARY
4.1.2.48
-
RECOMMENDED NAME
GeneOntology No.
low-specificity L-threonine aldolase
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
L-allo-threonine = glycine + acetaldehyde
show the reaction diagram
-
-
-
-
L-threonine = glycine + acetaldehyde
show the reaction diagram
(1)
-
-
-
PATHWAY
KEGG Link
MetaCyc Link
glycine biosynthesis IV
-
threonine degradation IV
-
SYSTEMATIC NAME
IUBMB Comments
L-threonine/L-allo-threonine acetaldehyde-lyase (glycine-forming)
Requires pyridoxal phosphate. The low-specificity L-threonine aldolase can act on both L-threonine and L-allo-threonine [1,2]. The enzyme from Escherichia coli can also act on L-threo-phenylserine and L-erythro-phenylserine [4]. The enzyme can also catalyse the aldol condensation of glycolaldehyde and glycine to form 4-hydroxy-L-threonine, an intermediate of pyridoxal phosphate biosynthesis [3]. Different from EC 4.1.2.5, L-threonine aldolase, and EC 4.1.2.49, L-allo-threonine aldolase.
SYNONYMS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
GLY1
P37303
gene name
GlyA
P0A825
-
L-TA
O50584
-
L-TA
O50584
-
-
L-threonine aldolase
-
-
L-threonine aldolase
-
-
low specificity L-TA
P37303
-
LtaE
-
-
-
-
LtaE
P75823
-
serine hydroxy-methyl transferase
-
-
threonine aldolase
-
-
ORGANISM
COMMENTARY
LITERATURE
SEQUENCE CODE
SEQUENCE DB
SOURCE
strain SGY269
SwissProt
Manually annotated by BRENDA team
Candida albicans SGY269
strain SGY269
SwissProt
Manually annotated by BRENDA team
Caulobacter vibrioides CB15
-
UniProt
Manually annotated by BRENDA team
serine hydroxymethyl transferase with threonine aldolase activity; MG1655
Uniprot
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
malfunction
-, P75823
knockout of the ltaE gene of wild-type Escherichia coli does not affect the cellular growth rate, while disruption of the ltaE gene of Escherichia coli GS245, whose serine hydroxymethyltransferase gene is knocked out, causes a significant decrease in the cellular growth rate, suggesting that the threonine aldolase is not a major source of cellular glycine in wild-type Escherichia coli but catalyzes an alternative pathway for cellular glycine when serine hydroxymethyltransferase is inert
physiological function
-, P75823
threonine aldolase is not a major source of cellular glycine in wild-type Escherichia coli but catalyzes an alternative pathway for cellular glycine when serine hydroxymethyltransferase is inert
physiological function
-
low-specificity L-threonine aldolase is involved in a serendipitous pathway that converts 3-phosphohydroxypyruvate, an intermediate in the serine biosynthesis pathway, to L-4-phosphohydroxythreonine, an intermediate in the pyridoxal-5'-phosphate synthesis pathway in a strain of Escherichia coli that lacks 4-phosphoerythronate dehydrogenase
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
3,4-dihydroxybenzaldehyde + glycine
(2S,3R,4R)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 40%, glycine concentration: 70 mM, reaction temperature: 4C, yield: 30%, L-erythro/L-threo: 16:84
-
-
?
3,4-dihydroxybenzaldehyde + glycine
(2S,3R,4R)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 60%, glycine concentration: 70 mM, reaction temperature: 4C, yield: 30%, L-erythro/L-threo: 100:0
-
-
?
3,4-dihydroxybenzaldehyde + glycine
(2S,3S,4R)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 40%, glycine concentration: 70 mM, reaction temperature: 4C, yield: 30%, L-erythro/L-threo: 16:84
-
-
?
benzyloxyacetaldehyde + glycine
(2S,3R)-2-amino-4-(benzyloxy)-3-hydroxybutanoic acid
show the reaction diagram
-
conversion: 45%, glycine concentration: 140 mM, reaction temperature: 25C, yield: 30%, L-erythro/L-threo: 40:60
-
-
?
benzyloxyacetaldehyde + glycine
(2S,3R)-2-amino-4-(benzyloxy)-3-hydroxybutanoic acid
show the reaction diagram
-
conversion: 68%, glycine concentration: 70 mM, reaction temperature: 4C, yield: 40%, L-erythro/L-threo: 97:3
-
-
?
benzyloxyacetaldehyde + glycine
(2S,3S)-2-amino-4-(benzyloxy)-3-hydroxybutanoic acid
show the reaction diagram
-
conversion: 45%, glycine concentration: 140 mM, reaction temperature: 25C, yield: 30%, L-erythro/L-threo: 40:60
-
-
?
DL-erythro-phenylserine
glycine + benzaldehyde
show the reaction diagram
-, P75823
-
-
-
?
DL-threo-(3-methylsulfonylphenyl)serine
glycine + 3-methylsulfonylbenzaldehyde
show the reaction diagram
P0A825
121% of the activity with L-threonine
-
-
?
DL-threo-(3-nitrophenyl)serine
glycine + 3-nitrobenzaldehyde
show the reaction diagram
P0A825
143% of the activity with L-threonine
-
-
?
DL-threo-phenylserine
glycine + benzaldehyde
show the reaction diagram
-, P75823
-
-
-
?
DL-threo-phenylserine
glycine + benzaldehyde
show the reaction diagram
P0A825
180% of the activity with L-threonine
-
-
?
glycine + 3,4-dihydroxybenzaldehyde
L-threo-3,4-dihydroxyphenylserine
show the reaction diagram
-
-
-
-
r
glycine + 3,4-dihydroxybenzaldehyde
L-threo-3,4-dihydroxyphenylserine
show the reaction diagram
-
L-threo-3,4-dihydroxyphenylserine synthesis activity is dramatically decreased when the condensation reaction is repeated
-
-
-
glycine + 3,4-dihydroxybenzaldehyde
L-threo-3,4-dihydroxyphenylserine + L-erythro-3,4-dihydroxyphenylserine
show the reaction diagram
-
-
-
-
r
glycine + glycolaldehyde
L-4-hydroxythreonine
show the reaction diagram
-
low-specificity L-threonine aldolase is involved in a serendipitous pathway that converts 3-phosphohydroxypyruvate, an intermediate in the serine biosynthesis pathway, to L-4-phosphohydroxythreonine, an intermediate in the pyridoxal-5'-phosphate synthesis pathway in a strain of Escherichia coli that lacks 4-phosphoerythronate dehydrogenase
-
-
r
L-4-hydroxythreonine
glycine + glycolaldehyde
show the reaction diagram
-
cleavage of L-4-hydroxythreonine is as efficient as cleavage of L-allo-threonine
-
-
r
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
-
-
-
-
r
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
-
-
-
-
r
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
P37303
-
-
-
?
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
-, P75823
-
-
-
r
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
O50584, -
-
-
-
r
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
Q9A3V8
-
-
-
?
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
E7U392
-
-
-
?
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
P0A825
291% of the activity with L-threonine
-
-
?
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
Caulobacter vibrioides CB15
Q9A3V8
-
-
-
?
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
E7U392
-
-
-
?
L-allo-threonine
glycine + acetaldehyde
show the reaction diagram
O50584
-
-
-
r
L-erythro-phenylserine
glycine + benzaldehyde
show the reaction diagram
O50584, -
-
-
-
?
L-erythro-phenylserine
glycine + benzaldehyde
show the reaction diagram
O50584
-
-
-
?
L-phenylserine
?
show the reaction diagram
-
-
-
-
?
L-serine
glycine + formaldehyde
show the reaction diagram
-
-
-
-
?
L-Thr
Gly + acetaldehyde
show the reaction diagram
-
-
-
-
?
L-threo-3,4-dihydroxyphenylserine
glycine + 3,4-dihydroxybenzaldehyde
show the reaction diagram
-
-
-
-
?
L-threo-beta-3,4-dihydroxyphenylserine
glycine + 3,4-dihydroxybenzaldehyde
show the reaction diagram
O50584, -
-
-
-
?
L-threo-beta-3,4-dihydroxyphenylserine
glycine + 3,4-dihydroxybenzaldehyde
show the reaction diagram
O50584
-
-
-
?
L-threo-beta-3,4-methylenedioxyphenylserine
?
show the reaction diagram
O50584, -
-
-
-
?
L-threo-beta-3,4-methylenedioxyphenylserine
?
show the reaction diagram
O50584
-
-
-
?
L-threo-phenylserine
glycine + benzaldehyde
show the reaction diagram
O50584, -
-
-
-
?
L-threonine
glycine + acetaldehyde
show the reaction diagram
-
-
-
-
r
L-threonine
glycine + acetaldehyde
show the reaction diagram
-
-
-
-
r
L-threonine
glycine + acetaldehyde
show the reaction diagram
-, O13427
-
-
r
L-threonine
glycine + acetaldehyde
show the reaction diagram
P37303
-
-
-
r
L-threonine
glycine + acetaldehyde
show the reaction diagram
-, P75823
-
-
-
r
L-threonine
glycine + acetaldehyde
show the reaction diagram
O50584, -
-
-
-
r
L-threonine
glycine + acetaldehyde
show the reaction diagram
P0A825
-
-
-
?
L-threonine
glycine + acetaldehyde
show the reaction diagram
Q9A3V8
-
-
-
?
L-threonine
glycine + acetaldehyde
show the reaction diagram
E7U392
-
-
-
?
L-threonine
glycine + acetaldehyde
show the reaction diagram
Caulobacter vibrioides CB15
Q9A3V8
-
-
-
?
L-threonine
glycine + acetaldehyde
show the reaction diagram
Candida albicans SGY269
O13427
-
-
r
L-threonine
glycine + acetaldehyde
show the reaction diagram
E7U392
-
-
-
?
N-(S)-benzyloxycarbonyl-alaninal + glycine
(2S,3R,4S)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 48%, glycine concentration: 280 mM, reaction temperature: 4C, yield: 30%, L-erythro/L-threo: 100:0
-
-
?
N-(S)-benzyloxycarbonyl-alaninal + glycine
(2S,3R,4S)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 54%, glycine concentration: 140 mM, reaction temperature: 25C, yield: 27%, L-erythro/L-threo: 18:82
-
-
?
N-(S)-benzyloxycarbonyl-alaninal + glycine
(2S,3S,4S)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 54%, glycine concentration: 140 mM, reaction temperature: 25C, yield: 27%, L-erythro/L-threo: 18:82
-
-
?
N-benzyloxycarbonyl-3-aminopropanal + glycine
(2S,3R)-2-amino-5-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 34%, glycine concentration: 70 mM, reaction temperature: 25C, yield: 10%, L-erythro/L-threo: 50:50
-
-
?
N-benzyloxycarbonyl-3-aminopropanal + glycine
(2S,3R)-2-amino-5-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 49%, glycine concentration: 70 mM, reaction temperature: 25C, yield: 11%, L-erythro/L-threo: 50:50
-
-
?
N-benzyloxycarbonyl-3-aminopropanal + glycine
(2S,3S)-2-amino-5-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 20%, glycine concentration: 70 mM, reaction temperature: 4C, yield: 3%, L-erythro/L-threo: 100:0
-
-
?
N-benzyloxycarbonyl-3-aminopropanal + glycine
(2S,3S)-2-amino-5-(benzyloxycarbonylamino)-3-hydroxypentanoic acid
show the reaction diagram
-
conversion: 49%, glycine concentration: 70 mM, reaction temperature: 25C, yield: 11%, L-erythro/L-threo: 50:50
-
-
?
N-benzyloxycarbonyl-glycinal + glycine
(2S,3R)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxybutanoic acid
show the reaction diagram
-
conversion: 35%, glycine concentration: 70 mM, reaction temperature: 4C, yield: 13%, L-erythro/L-threo: 86:14
-
-
?
N-benzyloxycarbonyl-glycinal + glycine
(2S,3R)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxybutanoic acid
show the reaction diagram
-
conversion: 60%, glycine concentration: 140 mM, reaction temperature: 25C, yield: 18%, L-erythro/L-threo: 30:70
-
-
?
N-benzyloxycarbonyl-glycinal + glycine
(2S,3S)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxybutanoic acid
show the reaction diagram
-
conversion: 35%, glycine concentration: 70 mM, reaction temperature: 4C, yield: 13%, L-erythro/L-threo: 86:14
-
-
?
N-benzyloxycarbonyl-glycinal + glycine
(2S,3S)-2-amino-4-(benzyloxycarbonylamino)-3-hydroxybutanoic acid
show the reaction diagram
-
conversion: 60%, glycine concentration: 140 mM, reaction temperature: 25C, yield: 18%, L-erythro/L-threo: 30:70
-
-
?
L-threonine
glycine + acetaldehyde
show the reaction diagram
O50584
-
-
-
r
additional information
?
-
-, O13427
glycine metabolism
-
?
additional information
?
-
-
by manipulating reaction parameters, SHMT yields exclusively L-erythro diastereomers in 34-60% conversion. SHMT is among the most stereoselective L-threonine aldolases described. This is due to its activity-temperature dependence: at 4C SHMT has high synthetic activity but negligible retro-aldol activity on l-threonine. Thus, the kinetic l-erythro isomer is largely favored and the reactions are virtually irreversible, highly stereoselective, and in turn, give excellent conversion
-
-
-
additional information
?
-
Candida albicans SGY269
O13427
glycine metabolism
-
?
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
glycine + glycolaldehyde
L-4-hydroxythreonine
show the reaction diagram
-
low-specificity L-threonine aldolase is involved in a serendipitous pathway that converts 3-phosphohydroxypyruvate, an intermediate in the serine biosynthesis pathway, to L-4-phosphohydroxythreonine, an intermediate in the pyridoxal-5'-phosphate synthesis pathway in a strain of Escherichia coli that lacks 4-phosphoerythronate dehydrogenase
-
-
r
L-Thr
Gly + acetaldehyde
show the reaction diagram
-
-
-
-
?
additional information
?
-
-, O13427
glycine metabolism
-
?
additional information
?
-
Candida albicans SGY269
O13427
glycine metabolism
-
?
COFACTOR
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
pyridoxal 5'-phosphate
O50584, -
contains 1 mol of pyridoxal 5'-phosphate per mol of 38000 da subunit. Lys207 probably functions as an essential catalytic residue, forming an internal Schiff base with the pyridoxal 5'-phosphate of the enzyme to catalyze the reversible aldol reaction
pyridoxal 5'-phosphate
-
Km: 0.00025 mM, 1 mol of pyridoxal phosphate binds 46000 g of protein
pyridoxal 5'-phosphate
-
contains 6 mol of pyridoxal 5'-phosphate per mol of enzyme
pyridoxal 5'-phosphate
P37303
2 mol pyridoxal 5'-phosphate per 4 mol of subunit
pyridoxal 5'-phosphate
-, P75823
contains 1 mol of pyridoxal 5'-phosphate as cofactor per mol of 36500 Da subunit
pyridoxal 5'-phosphate
-
activates
pyridoxal 5'-phosphate
-
-
METALS and IONS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
K+
-
K+ or NH4+ required for maximal activity
INHIBITORS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
Ba2+
-
inhibited activation by K+ or NH4+
Ca2+
-
inhibited activation by K+ or NH4+
Mg2+
-
inhibited activation by K+ or NH4+
Na+
-
inhibited activation by K+ or NH4+
tetrahydrofolate
-
inhibits interconversion of L-serine and glycine
ACTIVATING COMPOUND
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
NH4+
-
K+ or NH4+ required for maximal activity
sodium sulfite
A0T1V9
addition of sodium sulfite stimulates the production of L-threo-3,4-dihydroxyphenylserine without affecting the diastereoselectivity ratio, especially at 50 mM
Triton X-100
A0T1V9
highest conversion yield at a 0.75% without affecting the diastereoselectivity ratio
KM VALUE [mM]
KM VALUE [mM] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.24
-
DL-erythro-phenylserine
-, P75823
pH 8.0, 30C
23.6
-
DL-threo-(3-methylsulfonylphenyl)serine
P0A825
pH 7.0, 37C
-
21.5
-
DL-threo-(3-nitrophenyl)serine
P0A825
pH 7.0, 37C
-
0.12
-
DL-threo-phenylserine
-, P75823
pH 8.0, 30C
19.2
-
DL-threo-phenylserine
P0A825
pH 7.0, 37C
0.027
-
L-4-hydroxythreonine
-
pH 8.0, 25C
-
0.052
-
L-allo-threonine
-
pH 8.0, 25C
0.2
-
L-allo-threonine
E7U392
mutant H126NF, pH 7.0, 30C
0.22
-
L-allo-threonine
-, P75823
pH 8.0, 30C
0.24
-
L-allo-threonine
E7U392
wild-type, pH 7.0, 30C
0.69
-
L-allo-threonine
Q9A3V8
wild-type, pH 8.0, 30C
0.96
-
L-allo-threonine
E7U392
mutant H126N, pH 7.0, 30C
1
2
L-allo-threonine
Q9A3V8
mutant D95L, pH 8.0, 30C
1.3
-
L-allo-threonine
E7U392
mutant F87A, pH 7.0, 30C
1.5
-
L-allo-threonine
P0A825
pH 7.0, 37C
1.7
-
L-allo-threonine
E7U392
mutant F87D, pH 7.0, 30C; mutant H83N, pH 7.0, 30C
2.5
-
L-allo-threonine
Q9A3V8
mutant D95W, pH 8.0, 30C
2.7
-
L-allo-threonine
Q9A3V8
mutant D95Y, pH 8.0, 30C
2.9
-
L-allo-threonine
Q9A3V8
mutant D95G/E96G, pH 8.0, 30C
4.4
-
L-allo-threonine
Q9A3V8
mutant D95Y/E96T, pH 8.0, 30C
5
-
L-allo-threonine
E7U392
mutant K222A, pH 7.0, 30C
7
-
L-allo-threonine
E7U392
mutant H83F, pH 7.0, 30C
7.1
-
L-allo-threonine
Q9A3V8
mutant D95N/E96S, pH 8.0, 30C
10
-
L-allo-threonine
P37303
pH 8.0, 30C
11
-
L-allo-threonine
Q9A3V8
mutant D95C, pH 8.0, 30C
14.6
-
L-allo-threonine
O50584, -
pH 8.0, 30C
31
-
L-allo-threonine
E7U392
mutant H83F/H126F, pH 7.0, 30C
10.2
-
L-erythro-phenylserine
O50584, -
pH 8.0, 30C
0.000002
-
L-phenylserine
-
recombinant wild type enzyme, in 50 mM Tris-HCl buffer (pH 8.0)
0.0000026
-
L-phenylserine
-
mutant enzyme H177Y, in 50 mM Tris-HCl buffer (pH 8.0)
0.00016
-
L-Thr
-
mutant enzyme H177Y, in 50 mM Tris-HCl buffer (pH 8.0)
0.00018
-
L-Thr
-
recombinant wild type enzyme, in 50 mM Tris-HCl buffer (pH 8.0)
0.0002
-
L-threo-3,4-dihydroxyphenylserine
-
recombinant wild type enzyme, in 50 mM Tris-HCl buffer (pH 8.0)
0.00023
-
L-threo-3,4-dihydroxyphenylserine
-
mutant enzyme H177Y, in 50 mM Tris-HCl buffer (pH 8.0)
8.3
-
L-threo-beta-3,4-dihydroxyphenylserine
O50584, -
pH 8.0, 30C
7.4
-
L-threo-beta-3,4-methylenedioxyphenylserine
O50584, -
pH 8.0, 30C
7.3
-
L-threo-phenylserine
O50584, -
pH 8.0, 30C
0.4
-
L-threonine
E7U392
mutant F87A, pH 7.0, 30C
1
3
L-threonine
Q9A3V8
mutant D95G/E96G, pH 8.0, 30C
1.1
-
L-threonine
E7U392
mutant H83F/H126F, pH 7.0, 30C
1.3
-
L-threonine
E7U392
mutant F87D, pH 7.0, 30C
1.7
-
L-threonine
E7U392
mutant H126F, pH 7.0, 30C
2.85
-
L-threonine
-, P75823
pH 8.0, 30C
3.7
-
L-threonine
E7U392
mutant K222A, pH 7.0, 30C
4
-
L-threonine
-
pH 8.0, 25C
7
-
L-threonine
E7U392
mutant H83F, pH 7.0, 30C
13.6
-
L-threonine
Q9A3V8
wild-type, pH 8.0, 30C
14.7
-
L-threonine
O50584, -
pH 8.0, 30C
19.4
-
L-threonine
E7U392
wild-type, pH 7.0, 30C
32
-
L-threonine
Q9A3V8
mutant D95N/E96S, pH 8.0, 30C
38
-
L-threonine
E7U392
mutant H83N, pH 7.0, 30C
43.1
-
L-threonine
P0A825
pH 7.0, 37C
44
-
L-threonine
Q9A3V8
mutant D95C, pH 8.0, 30C; mutant D95W, pH 8.0, 30C
50
-
L-threonine
Q9A3V8
mutant D95M, pH 8.0, 30C
53
-
L-threonine
Q9A3V8
mutant D95L, pH 8.0, 30C
55
-
L-threonine
P37303
pH 8.0, 30C
61
-
L-threonine
E7U392
mutant H126N, pH 7.0, 30C
63
-
L-threonine
Q9A3V8
mutant D95Y, pH 8.0, 30C
TURNOVER NUMBER [1/s]
TURNOVER NUMBER MAXIMUM[1/s]
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
1.44
-
L-4-hydroxythreonine
-
pH 8.0, 25C
-
0.06
-
L-allo-threonine
E7U392
mutant H83F, pH 7.0, 30C
0.2
-
L-allo-threonine
Q9A3V8
mutant D95N/E96S, pH 8.0, 30C
1.28
-
L-allo-threonine
E7U392
mutant H83N, pH 7.0, 30C
3
-
L-allo-threonine
Q9A3V8
mutant D95Y/E96T, pH 8.0, 30C
3.2
-
L-allo-threonine
-
pH 8.0, 25C
3.55
-
L-allo-threonine
E7U392
wild-type, pH 7.0, 30C
7.82
-
L-allo-threonine
E7U392
mutant H126N, pH 7.0, 30C
8.6
-
L-allo-threonine
Q9A3V8
mutant D95G/E96G, pH 8.0, 30C
9.02
-
L-allo-threonine
E7U392
mutant H126NF, pH 7.0, 30C
19
-
L-allo-threonine
Q9A3V8
mutant D95C, pH 8.0, 30C; mutant D95W, pH 8.0, 30C
20
-
L-allo-threonine
Q9A3V8
mutant D95Y, pH 8.0, 30C; wild-type, pH 8.0, 30C
22
-
L-allo-threonine
Q9A3V8
mutant D95L, pH 8.0, 30C
41
-
L-allo-threonine
P37303
pH 8.0, 30C
0.000107
-
L-phenylserine
-
recombinant wild type enzyme, in 50 mM Tris-HCl buffer (pH 8.0)
0.000113
-
L-phenylserine
-
mutant enzyme H177Y, in 50 mM Tris-HCl buffer (pH 8.0)
0.0178
-
L-Thr
-
mutant enzyme H177Y, in 50 mM Tris-HCl buffer (pH 8.0)
0.0195
-
L-Thr
-
recombinant wild type enzyme, in 50 mM Tris-HCl buffer (pH 8.0)
0.00022
-
L-threo-3,4-dihydroxyphenylserine
-
recombinant wild type enzyme, in 50 mM Tris-HCl buffer (pH 8.0)
0.00023
-
L-threo-3,4-dihydroxyphenylserine
-
mutant enzyme H177Y, in 50 mM Tris-HCl buffer (pH 8.0)
0.02
-
L-threonine
E7U392
mutant H83F, pH 7.0, 30C
0.2
-
L-threonine
Q9A3V8
mutant D95N/E96S, pH 8.0, 30C
0.28
-
L-threonine
E7U392
mutant H83N, pH 7.0, 30C
0.6
-
L-threonine
Q9A3V8
mutant D95G/E96G, pH 8.0, 30C
1.1
-
L-threonine
-
pH 8.0, 25C
1.87
-
L-threonine
E7U392
wild-type, pH 7.0, 30C
3
-
L-threonine
Q9A3V8
mutant D95C, pH 8.0, 30C
3.3
-
L-threonine
Q9A3V8
wild-type, pH 8.0, 30C
3.5
-
L-threonine
Q9A3V8
mutant D95Y, pH 8.0, 30C
3.6
-
L-threonine
Q9A3V8
mutant D95M, pH 8.0, 30C
3.8
-
L-threonine
Q9A3V8
mutant D95L, pH 8.0, 30C
4.37
-
L-threonine
E7U392
mutant H126N, pH 7.0, 30C
6
-
L-threonine
E7U392
mutant H126F, pH 7.0, 30C
9.6
-
L-threonine
Q9A3V8
mutant D95W, pH 8.0, 30C
43
-
L-threonine
P37303
pH 8.0, 30C
kcat/KM VALUE [1/mMs-1]
kcat/KM VALUE [1/mMs-1] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
5.3
-
L-4-hydroxythreonine
-
pH 8.0, 25C
0
0.01
-
L-allo-threonine
E7U392
mutant H83F, pH 7.0, 30C
2967
0.028
-
L-allo-threonine
Q9A3V8
mutant D95N/E96S, pH 8.0, 30C
2967
0.68
-
L-allo-threonine
Q9A3V8
mutant D95Y/E96T, pH 8.0, 30C
2967
0.75
-
L-allo-threonine
E7U392
mutant H83N, pH 7.0, 30C
2967
1.7
-
L-allo-threonine
Q9A3V8
mutant D95C, pH 8.0, 30C
2967
1.8
-
L-allo-threonine
Q9A3V8
mutant D95L, pH 8.0, 30C
2967
3
-
L-allo-threonine
Q9A3V8
mutant D95G/E96G, pH 8.0, 30C
2967
4.1
-
L-allo-threonine
P37303
pH 8.0, 30C
2967
6.2
-
L-allo-threonine
-
pH 8.0, 25C
2967
7.4
-
L-allo-threonine
Q9A3V8
mutant D95Y, pH 8.0, 30C
2967
7.6
-
L-allo-threonine
Q9A3V8
mutant D95W, pH 8.0, 30C
2967
8.13
-
L-allo-threonine
E7U392
mutant H126N, pH 7.0, 30C
2967
14.78
-
L-allo-threonine
E7U392
wild-type, pH 7.0, 30C
2967
29
-
L-allo-threonine
Q9A3V8
wild-type, pH 8.0, 30C
2967
45.08
-
L-allo-threonine
E7U392
mutant H126NF, pH 7.0, 30C
2967
0.0016
-
L-threonine
Q9A3V8
mutant D95Y/E96T, pH 8.0, 30C
250
0.0063
-
L-threonine
Q9A3V8
mutant D95N/E96S, pH 8.0, 30C
250
0.01
-
L-threonine
E7U392
mutant H83N, pH 7.0, 30C
250
0.046
-
L-threonine
Q9A3V8
mutant D95G/E96G, pH 8.0, 30C
250
0.056
-
L-threonine
Q9A3V8
mutant D95Y, pH 8.0, 30C
250
0.068
-
L-threonine
Q9A3V8
mutant D95C, pH 8.0, 30C
250
0.07
-
L-threonine
E7U392
mutant H126N, pH 7.0, 30C
250
0.072
-
L-threonine
Q9A3V8
mutant D95L, pH 8.0, 30C; mutant D95M, pH 8.0, 30C
250
0.1
-
L-threonine
E7U392
wild-type, pH 7.0, 30C
250
0.218
-
L-threonine
Q9A3V8
mutant D95W, pH 8.0, 30C
250
0.24
-
L-threonine
Q9A3V8
wild-type, pH 8.0, 30C
250
0.78
-
L-threonine
P37303
pH 8.0, 30C
250
2.8
-
L-threonine
-
pH 8.0, 25C
250
3.53
-
L-threonine
E7U392
mutant H126F, pH 7.0, 30C
250
SPECIFIC ACTIVITY [µmol/min/mg]
SPECIFIC ACTIVITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
1.7
-
-, P75823
pH 8.0, 30C
2.5
-
P37303
pH 8.0, 30C
2.91
-
-
pH 8.5, 30C
2.91
-
-
pH 8.6, 30C
8.5
-
-, O13427
cell-free extract
41
-
O50584, -
pH 8.0, 30C
pH OPTIMUM
pH MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
6.5
-
A0T1V9
-
7
8
-
recombinant enzyme
8
8.5
O50584, -
-
8
-
O50584, -
assay at
8
-
P37303
assay at
8
-
-, P75823
assay at
8
-
-
assay at
8.5
9
P37303
aldehyde formation from L-allo-threonine
8.5
9
-, P75823
-
8.5
-
-
assay at
8.5
-
-
substrate: L-threonine, Tris-chloride buffer
10
-
-
substrate: L-allo-threonine, Tris-chloride buffer
TEMPERATURE OPTIMUM
TEMPERATURE OPTIMUM MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
25
-
O50584, -
-
25
-
-
assay at
30
-
O50584, -
assay at
30
-
-
assay at
30
-
P37303
assay at
30
-
-, P75823
assay at
50
-
-
recombinant enzyme
55
-
P37303
aldehyde formation from L-allo-threonine
65
70
-, P75823
-
MOLECULAR WEIGHT
MOLECULAR WEIGHT MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
37000
-
-
subunit, SDS-PAGE
41810
-
-, O13427
calculated from amino acid sequence
140000
-
-, P75823
gel filtration
145000
-
O50584, -
gel filtration
150000
-
-
gel filtration
170000
-
P37303
gel filtration
277000
-
-
ultracentrifugation
SUBUNITS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
homotetramer
P37303
4 * 41000, SDS-PAGE; 4 * 42000, calculated from sequence
homotetramer
-
4 * 37500, gel filtration
tetramer
O50584, -
4 * 38000, SDS-PAGE
tetramer
-, P75823
4 * 36500, SDS-PAGE
tetramer
-
4 * 38000, SDS-PAGE
-
Crystallization/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
at 2.2 A resolution, in the unliganded form and cocrystallized with L-serine and L-threonine. No active site catalytic residue is revealed, and a structural water molecule is assumed to act as the catalytic base in the retro-aldol cleavage reaction. The very large active site opening suggests that much larger molecules than L-threonine isomers may be easily accommodated
E7U392
pH STABILITY
pH STABILITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
5.5
9
O50584, -
30C, 30 min, stable
6
9.5
-, P75823
30C, 30 min, stable
6.5
10
P37303
30 min, stable
TEMPERATURE STABILITY
TEMPERATURE STABILITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
25
-
O50584, -
40C, 15 min, 50% loss of activity
50
-
P37303
15 min, enzyme retains only 25% of activity
60
-
-, P75823
1 h, stable
60
-
-
the residual L-TA activity after a heat treatment for 20 min at 60C is 10.6%
63
-
-
the half-life of the wild type L-TA at 63C is 1.3 min
Purification/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
-
-, P75823
-
O50584, -
Hiprep 16/10 DEAE FF column chromatography, Super Sepharose column chromatography, and HiLoad 16/10 Phenyl Sepharose HP column chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
expression in Escherichia coli
E7U392
overexpression in Escherichia coli
O50584, -
cloned into pUCl18, and expressed in Escherichia coli
P37303
expression in Escherichia coli
A0T1V9
recombinantly expressed in Escherichia coli
-
expressed in Escherichia coli strain JM109
-
ENGINEERING
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
D176E
Q9A3V8
500fold decrease in catalytic efficiency
D95C
Q9A3V8
less than 10% of catalytic efficiency of wild-type
D95H/E96G
Q9A3V8
less than 10% of catalytic efficiency of wild-type
D95L
Q9A3V8
less than 10% of catalytic efficiency of wild-type
D95M
Q9A3V8
less than 5% of catalytic efficiency of wild-type
D95N/E96S
Q9A3V8
less than 5% of catalytic efficiency of wild-type
D95W
Q9A3V8
less than 2% of catalytic efficiency of wild-type
D95Y
Q9A3V8
less than 5% of catalytic efficiency of wild-type
D95Y/E96T
Q9A3V8
less than 10% of catalytic efficiency of wild-type
D176E
Caulobacter vibrioides CB15
-
500fold decrease in catalytic efficiency
-
D95C
Caulobacter vibrioides CB15
-
less than 10% of catalytic efficiency of wild-type
-
D95L
Caulobacter vibrioides CB15
-
less than 10% of catalytic efficiency of wild-type
-
D95M
Caulobacter vibrioides CB15
-
less than 5% of catalytic efficiency of wild-type
-
D95Y
Caulobacter vibrioides CB15
-
less than 5% of catalytic efficiency of wild-type
-
F87A
E7U392
no change in the ration of cleavage of L-threonine to L-allo-threonine
F87D
E7U392
mutation doubles the preference of the enzyme for L-allo-threonine
H126F
E7U392
300% of wild-type activity,reduced preference for the erythro-substrate
H126N
E7U392
60% of wild-type activity
H83F
E7U392
less than 1% of wild-type activity, reduced preference for the erythro-substrate
H83F/H126F
E7U392
able to catalyze the cleavage of both L-threonine and L-allo-threonine at a measurable rate, neither of the histidines acts as a catalytic base in the retro-aldol cleavage mechanism
H83N
E7U392
less than 10% of wild-type activity
K222A
E7U392
slight decrease in kcat and slight increase in Km values for both L-threonine and L-allo-threonine
F87A
-
no change in the ration of cleavage of L-threonine to L-allo-threonine
-
F87D
-
mutation doubles the preference of the enzyme for L-allo-threonine
-
H126F
-
300% of wild-type activity,reduced preference for the erythro-substrate
-
H126N
-
60% of wild-type activity
-
K222A
-
slight decrease in kcat and slight increase in Km values for both L-threonine and L-allo-threonine
-
K207A
O50584, -
the mutant enzyme shows no detectable enzyme activity. The mutant enzyme show the disappearance of the absorption maximum at 420 nm, indicating that the Schiff base linkage between the epsilon-amino group of the active-site lysine residue and the pyridoxal 5'-phosphate cofactor aldehyde group of the wild type is not present in the mutant enzyme
K207R
O50584, -
the mutant enzyme shows a specific activity of about 1000 times lower than that of the wild-type enzyme. The mutant enzyme show the disappearance of the absorption maximum at 420 nm, indicating that the Schiff base linkage between the epsilon-amino group of the active-site lysine residue and the pyridoxal 5'-phosphate cofactor aldehyde group of the wild type is not present in the mutant enzyme
K207A
-
the mutant enzyme shows no detectable enzyme activity. The mutant enzyme show the disappearance of the absorption maximum at 420 nm, indicating that the Schiff base linkage between the epsilon-amino group of the active-site lysine residue and the pyridoxal 5'-phosphate cofactor aldehyde group of the wild type is not present in the mutant enzyme
-
K207R
-
the mutant enzyme shows a specific activity of about 1000 times lower than that of the wild-type enzyme. The mutant enzyme show the disappearance of the absorption maximum at 420 nm, indicating that the Schiff base linkage between the epsilon-amino group of the active-site lysine residue and the pyridoxal 5'-phosphate cofactor aldehyde group of the wild type is not present in the mutant enzyme
-
A169T
-
stability-enhanced mutant, half life at 63C is 3.7 min
D104N
-
stability-enhanced mutant, half life at 63C is 5.8 min
F18I
-
stability-enhanced mutant, half life at 63C is 5.0 min, the specific activity is decreased by 45% compared to the wild type enzyme
H177Y
-
stability-enhanced mutant, half life at 63C is 14.6 min
R241C/A287V
-
the mutations dramatically increase the diastereoselectivity of the reverse aldol condensation activity for L-threo-3,4-dihydroxyphenylserine
V86I/R241C/Y306C
-
the mutations dramatically increase the diastereoselectivity of the reverse aldol condensation activity for L-threo-3,4-dihydroxyphenylserine
Y34C
-
the mutation dramatically increases the diastereoselectivity of the reverse aldol condensation activity for L-threo-3,4-dihydroxyphenylserine
Y39C/Y306C
-
the mutations dramatically increase the diastereoselectivity of the reverse aldol condensation activity for L-threo-3,4-dihydroxyphenylserine
Y39C/Y306C/A48T
-
the mutations dramatically increase the diastereoselectivity of the reverse aldol condensation activity for L-threo-3,4-dihydroxyphenylserine
Y39C/Y306C/R316C
-
the mutations dramatically increase the diastereoselectivity of the reverse aldol condensation activity for L-threo-3,4-dihydroxyphenylserine
APPLICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
synthesis
P0A825
synthesis of optically active beta-hydroxy-alpha-amino acids by immobilized Escherichia coli cells expressing the enzyme. The immobilized cells can be continuously used 10 times, yielding an average conversion rate of 60.4%
synthesis
A0T1V9
production of L-threo-3,4-dihydroxyphenylserine. At the optimized conditions, a mixture of L-threo-3,4-dihydroxyphenylserine and L-erythro-3,4-dihydroxyphenylserine is synthesized by diastereoselectivity-enhanced L-threonine aldolase expressed in Escherichia coli in a continuous process for 100 h, yielding an average of 4.0 mg/ml of L-threo-3,4-dihydroxyphenylserine and 60% diastereoselectivity
biotechnology
-
a continuous bioconversion system for L-threo-3,4-dihydroxyphenylserine production is developed that uses whole-cell biocatalyst of recombinant Escherichia coli expressing L-TA genes cloned from Streptomyces avelmitilis MA-4680. Maximum conversion rates are observed at 2 M glycine, 145 mM 3,4-dihydroxybenzaldehyde, 0.75% Triton-X, 5 g Escherichia coli cells/l, pH 6.5 and 10C. In the optimized condition, overall productivity is 8 g/l