Information on EC 1.5.99.B2 - proline dehydrogenase (acceptor)

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

EC NUMBER
COMMENTARY hide
1.5.99.B2
preliminary BRENDA-supplied EC number
RECOMMENDED NAME
GeneOntology No.
proline dehydrogenase (acceptor)
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
L-proline + a quinone = (S)-1-pyrroline-5-carboxylate + a quinol
show the reaction diagram
L-proline + acceptor = (S)-1-pyrroline-5-carboxylate + reduced acceptor
show the reaction diagram
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
proline metabolism
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SYSTEMATIC NAME
IUBMB Comments
L-proline:acceptor oxidoreductase
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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-
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Manually annotated by BRENDA team
subspecies Bacillus subtilis natto
UniProt
Manually annotated by BRENDA team
tsetse fly
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Manually annotated by BRENDA team
squid
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Manually annotated by BRENDA team
11 different species
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Manually annotated by BRENDA team
MsPDH1; L.cv. Gilboa; MsPDH2; L. cv. Gilboa
SwissProt
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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SwissProt
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
corn
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Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
3,4-dehydro-L-proline + acceptor + H2O
? + reduced acceptor
show the reaction diagram
-
efficient substrate
-
-
?
4-methylene-L-proline + acceptor + H2O
DELTA1-pyrroline-3-methylene-5-carboxylic acid + reduced acceptor
show the reaction diagram
-
-
-
?
glycine + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
L-alanine + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
L-alanine + 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium + H2O
?
show the reaction diagram
L-arginine + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
L-histidine + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
L-hydroxyproline + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
L-hydroxyproline + 2,6-dichlorophenolindophenol + H2O
DELTA1-pyrroline-5-carboxylate + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
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100% activity compared to L-proline
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-
?
L-hydroxyproline + oxidized 2,6-dichlorophenolindophenol + H2O
?
show the reaction diagram
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reaction rate is 39% of that with L-proline
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-
?
L-leucine + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
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42% activity compared to L-proline
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-
?
L-pipecolic acid + acceptor + H2O
? + reduced acceptor
show the reaction diagram
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-
-
-
?
L-proline + 2,6-dichlorophenolindophenol + H2O
(S)-1-pyrroline-5-carboxylate + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
L-proline + 2,6-dichlorophenolindophenol + H2O
DELTA1-pyrroline-5-carboxylate + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
L-proline + 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium + H2O
DELTA1-pyrroline-5-carboxylate + reduced 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium
show the reaction diagram
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
show the reaction diagram
L-proline + coenzyme Q1 + H2O
DELTA1-pyrroline-5-carboxylate + reduced coenzyme Q1
show the reaction diagram
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-
-
?
L-proline + cytochrome c + H2O
(S)-1-pyrroline-5-carboxylate + reduced cytochrome c
show the reaction diagram
L-proline + FAD
(S)-1-pyrroline-5-carboxylate + FADH2
show the reaction diagram
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-
-
?
L-proline + ferricyanide
DELTA1-pyrroline-5-carboxylate + ferrocyanide
show the reaction diagram
L-proline + H2O
(S)-1-pyrroline-5-carboxylate + H2O2
show the reaction diagram
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-
-
-
?
L-proline + oxidized 2,6-dichlorophenolindophenol + H2O
(S)-1-pyrroline-5-carboxylate + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
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the enzyme does not utilize ferricyanide or 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide as the electron acceptor, activity with phenazine methosulfate-p-iodonitrotetrazolium violet is 10% of that with 2,6-dichlorophenolindophenol
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-
?
L-proline + p-iodonitrotetrazolium + H2O
(S)-1-pyrroline-5-carboxylate + reduced p-iodonitrotetrazolium
show the reaction diagram
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-
-
?
L-proline + phenazine methosulfate 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium
DELTA1-pyrroline-5-carboxylate + reduced phenazine methosulfate 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium
show the reaction diagram
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-
-
-
?
L-proline + phenazine methosulfate-4-iodonitrotetrazolium violet + H2O
DELTA1-pyrroline-5-carboxylate + reduced phenazine methosulfate-4-iodonitrotetrazolium violet
show the reaction diagram
L-serine + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
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55% activity compared to L-proline
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?
L-threonine + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
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66% activity compared to L-proline
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?
L-threonine + 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium + H2O
?
show the reaction diagram
L-valine + 2,6-dichlorophenolindophenol + H2O
? + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
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33% activity compared to L-proline
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?
sarcosine + acceptor + H2O
? + reduced acceptor
show the reaction diagram
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?
additional information
?
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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-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
HgCl2
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1 mM, 68% loss of activity
MgCl2
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1 mM, 50% loss of activity
additional information
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no significant effect on activity with 1 mM SrCl2, CaCl2, CoCl2, MgCl2, CuCl2, ZnCl2
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4-methylene-L-proline
abscisic acid
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L-Mandelate
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L-pyroglutamate
L-tetrahydro-2-furoic acid
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competitive inhibitor
Lactate
N-propargylglycine
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pyrrole-2-carboxylic acid
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noncompetitive
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
L-proline
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in the presence of L-proline, high POX activity is sufficient to induce mitochondria-mediated apoptosis
additional information
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POX mRNA expression, and the extent of POX induction is correlated with radical oxygen species level and cell death suggesting that POX-induced radical oxygen species formation is critical for the apoptotic cell death induced by PPARgamma activation
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.026 - 0.035
2,6-dichlorophenolindophenol
4
3,4-dehydro-L-proline
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in 50 mM Tris-HCl, at 25C
0.028 - 0.155
coenzyme Q1
6.05
L-hydroxyproline
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in 200 mM Tris-HCl buffer (pH 8.0), at 50C
212.3
L-Pipecolic acid
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wild type enzyme, in 100 mM potassium phosphate buffer, pH 7.5, at 80C
0.174 - 384
L-proline
0.0159
menadione
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0.14
oxidized 2,6-dichlorophenolindophenol
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pH 6.5, 50C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
75
3,4-dehydro-L-proline
Thermus thermophilus
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in 50 mM Tris-HCl, at 25C
0.043 - 14
coenzyme Q1
0.4
L-Pipecolic acid
Pyrococcus furiosus
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wild type enzyme in 100 mM potassium phosphate buffer, pH 7.5, at 80C
0.055 - 37.17
L-proline
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.9
D,L-lactate
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2.5
L-lactate
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in 50 mM Tris-HCl, at 25C
2.4
L-Mandelate
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in 50 mM Tris-HCl, at 25C
24 - 167
L-pyroglutamate
1
L-tetrahydro-2-furoic acid
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in 50 mM Tris-HCl, at 25C
0.2
N-propargylglycine
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at 25C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.026
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crude extract, pH 8.0, at 50C
0.939
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isoform PDH1
1.4
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isoform PDH2
3.57
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pH 6.5, 50C
4.25
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after 163fold purification, pH 8.0, at 50C
28.9
after purification, at 25?C, in 50 mM Tris-HCl (pH 7.5)
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.2
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proline + ferricyanide
7.5
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isoform PDH1
7.8
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L-proline + menadione
9.5
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L-proline + phenazine methosulfate
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 11
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more than 50% activity between pH 6.0 and 11.0
6 - 9
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half of the activity is retained at pH 6.0 and 9.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
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assay at
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 40
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more than 60% activity between 20-40C, a sharp decrease is observed above 30C and enzyme activity is completely inactivated at 70C
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
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isoelectric focusing
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
expression levels of ProDH2 are generally low, but increase in senescent leaves and in the abscission zone of floral organs
Manually annotated by BRENDA team
strongest expression in microspore; strongest expression in microspore
Manually annotated by BRENDA team
strong expression in mature pollen; strong expression in mature pollen
Manually annotated by BRENDA team
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during dark adaption PDH expression is significantly enhanced
Manually annotated by BRENDA team
weak expression; weak expression
Manually annotated by BRENDA team
additional information
expression levels of ProDH2 are generally low, but increase in senescent leaves and in the abscission zone of floral organs
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
34500
calculated from amino acid sequence
37970
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MALDI-TOF mass spectrometry
37980
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native enzyme, calculated from sequence of cDNA
42440
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beta subunit, MALDI-TOF mass spectrometry
42470
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beta subunit, electrospray ionization mass spectrometry
55000
SDS-PAGE; SDS-PAGE
94900
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gel filtration
101000
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isoform PDH2, gel filtration
108000
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gel filtration
242000
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gel filtration, native PAGE leads to two active forms of 24000 and 47000
440000
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isoform PDH1, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterooctamer
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x-ray crystallography
heterotetramer
homodimer
monomer
octamer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
enzyme bound with L-proline, sitting drop vapor diffusion method, using 0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate (pH 5.6), and 30% (w/v) polyethylene glycol 4000
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to 2.87 A resolution, space group P41212
His-tag removed, three crystal forms: apparent tetragonal, hexagonal and centered monoclinic
enzyme in the oxidized state complexed with the proline analogue L-tetrahydrofuroic acid and in the reduced state with the proline site vacant, sitting drop vapor diffusion method, using 0.2 M MgCl2, 25% (w/v) PEG 3350, and 0.1 mM Bis-Tris (pH 5.8), at 22C
beta subunit is the L-proline dehydrogenase catalytic component and contains FAD, alpha subunit has a classical dinucleotide fold domain with ATP and a Cys-clustered domain. FMN is located at the interface of alpha and beta subunits; crystal structure of PDH1, which is a heterooctameric complex containing three different cofactors: FAD, FMN, and ATP. The structure is determined by x-ray crystallography to a resolution of 2.86 A. The structure of the beta subunit, which is an L-proline dehydrogenase catalytic component containing FAD as a cofactor, is similar to that of monomeric sarcosine oxidase
from a monodisperse protein solution with detergent n-octyl beta-D-glucopyranoside
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sitting drop vapour diffusion method, with 100 mM imidazole (pH 7), 100 mM MgCl2, 17% 2-methyl-2,4-pentanediol, and 5 mM dithiothreitol
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sitting drop vapour diffusion method, with 100 mM imidazole buffer at pH 7, 100 mM MgCl2, 14% 2-methyl-2,4-pentanediol, and 5 mM dithiothreitol
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.5 - 9
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the enzyme retains more than 90% of its activity after incubation at pH values 4.5-9.0 for 30 min at 50C
724048
5 - 10
6 - 12
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50C, 30 min, stable
727195
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 70
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the enzyme loses a considerable amount of its original activity at above 30C and is nearly inactivated at 70C
45
the enzyme loses 70% of activity upon incubation at 45C for 15 min
80
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10 min, stable
100
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10 min, 48% loss of activity
100 - 105
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the enzyme retains more than 90% of its activity after incubation at 100C for 120 min (pH 7.5). The half-life at 105C is about 90 min
100
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no loss of activity being evident up to 100C, above this temperature, thermal denaturation of PRODH is apparent, with complete loss of activity after 10 min of incubation in glycerol buffer at temperatures above 115C
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, pH 7.6, 50% glycerol, 1 month
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
DE52 anion exchange column chromatography
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DEAE Toyopearl 650M column chromatography and Superdex 200 gel filtration
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HisTrap column chromatography and Hitrap Q column chromatography
Ni-chelating column chromatography
Ni-NTA column chromatography and Sephadex G-25 gel filtration
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Ni2+ affinity column chromatography
Ni2+ NTA affinity column chromatography
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Ni2+-charged chelating Sepharose column chromatography and Sephacryl S-300 gel filtration
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recombinant enzyme
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in DLD-1.POX cells
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expressed in Escherichia coli
expressed in Escherichia coli BL-21 (DE3) plysS cells
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expressed in Escherichia coli BL-21 plysS (DE3) cells
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expressed in Escherichia coli BL21 (DE3) cells
expressed in Escherichia coli BL21 (DE3) Codon Plus RILP cells
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expressed in Escherichia coli BL21 (DE3) pLysS cells
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expressed in Escherichia coli BL21(DE3) Codon Plus RIL cells
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expressed in Escherichia coli BL21(DE3)pLysS cells
expressed in Escherichia coli BL21-CodonPlus(DE3)-RIPL and BL21-Gold(DE3)pLysS cells
expressed in Escherichia coli Rosetta(DE3)pLysS cells
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expressed in H1299 cells
expression in Eshcerichia coli
N-terminal 8xHis tag
the two MsPDH genes MsPDH1 and MsPDH2 share a high nucleotide sequence homology and a similar exon/intron structure
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E64A
the mutation decreases the catalytic efficiency 27fold
G63A
the mutation decreases the catalytic efficiency 140fold
A167V
-
moderate reduction of POX activity
A455S
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slight reduction of POX activity
A472T
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slight reduction of POX activity
D426N
-
moderate reduction of POX activity
L289M
-
slight reduction of POX activity
L441P
-
severe reduction of POX activity
P460L
-
severe reduction of POX activity
Q19P
-
moderate reduction of POX activity
Q521E
-
severe reduction of POX activity
R185Q
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slight reduction of POX activity
R185W
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moderate reduction of POX activity
R431H
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moderate reduction of POX activity
R453C
-
severe reduction of POX activity
T466M
-
severe reduction of POX activity
V427M
-
moderate reduction of POX activity
H225A
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the mutant is unstable and precipitates from solution below pH 7.0, decreased activity compared to the wild type enzyme
H225Q
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the mutant displays activity down to solution pH 6.0, increased activity compared to the wild type enzyme
Y251F
-
wild type pH stability, increased activity compared to the wild type enzyme
additional information
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine
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proline dehydrogenase is involved in type I hyperprolinemia