BRENDA - Enzyme Database show
show all sequences of 1.14.14.10

Purification and characterization of a two-component monooxygenase that hydroxylates nitrilotriacetate from 'Chelatobacter' strain ATCC 29600

Uetz, T.; Schneider, R.; Snozzi, M.; Egli, T.; J. Bacteriol. 174, 1179-1188 (1992)

Data extracted from this reference:

Activating Compound
Activating Compound
Commentary
Organism
Structure
component A of NTA monooxygenase
the ability of component B of NTA monooxygenase to catalyze the oxidation of nitrilotriacetate to iminodiacetate and glyoxylate is completely dependent on the presence of component A
Aminobacter aminovorans
FMN
NTA-Mo activity increases by a factor of 4 when 0.003 mM FMN is included in the assay mixture, there is 0.4 mol of FMN per mol of purified component B
Aminobacter aminovorans
additional information
the activity is increased neither by additions of 20% (w/v) polyethylene glycol or sorbitol
Aminobacter aminovorans
Inhibitors
Inhibitors
Commentary
Organism
Structure
5,5'-dithiobis(2-nitrobenzoate)
67% residual activity at 1 mM
Aminobacter aminovorans
Ca2+
5% residual activity at 10 mM
Aminobacter aminovorans
EDTA
3% residual activity at 10 mM
Aminobacter aminovorans
FAD
24% residual activity at 0.1 mM
Aminobacter aminovorans
NaCl
54% residual activity at 250 mM
Aminobacter aminovorans
p-hydroxymercuribenzoate
12% residual activity at 1 mM
Aminobacter aminovorans
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.0013
-
FMN
in 40 mM TAPS buffer at pH 8.5 and 25C
Aminobacter aminovorans
0.5
-
nitrilotriacetate
in 40 mM TAPS buffer at pH 8.5 and 25C
Aminobacter aminovorans
Metals/Ions
Metals/Ions
Commentary
Organism
Structure
Co2+
may replace Mg2+, enzyme activity is reduced to 80% of that with MgCl2
Aminobacter aminovorans
Mg2+
required, 2 mM used in assay conditions
Aminobacter aminovorans
Mn2+
Mn2+ ions are able to replace Mg2+ but lead to a higher uncoupled NADH oxidation and enzyme activity is reduced to 70% of that with MgCl2
Aminobacter aminovorans
additional information
no nitrilotriacetate consumption is observed with Ca2+,Fe2+, Fe3+, Zn2+, Cu2+, or Ni2+; the enzyme contains less than 0.15 atom of Fe per mol of protein, indicating that neither Fe-sulfur clusters nor cytochromes are present
Aminobacter aminovorans
Molecular Weight [Da]
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
36000
-
2 * 36000, component B of NTA monooxygenase, SDS-PAGE
Aminobacter aminovorans
Organism
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
Aminobacter aminovorans
-
-
-
Aminobacter aminovorans ATCC 29600
-
-
-
Purification (Commentary)
Commentary
Organism
ammonium sulfate precipitation, phenyl Sepharose column chromatography, TMAE-Fractogel column chromatography, and phenyl Superose column chromatography
Aminobacter aminovorans
Storage Stability
Storage Stability
Organism
-70C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol, at pH 7.8, several months, without any significant loss of activity
Aminobacter aminovorans
4C, crude extract, 50 mM Tris-HCl, 200 h, 50% loss of activity
Aminobacter aminovorans
4C, purified component B of NTA monooxygenase in 20 mM HEPES buffer at pH 7.8, 7 days, 70% loss of activity
Aminobacter aminovorans
4C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol and ammonium sulfate to 5% saturation, at pH 7.8, 7 days, 5% loss of activity
Aminobacter aminovorans
4C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol, at pH 7.8, 7 days, 20% loss of activity
Aminobacter aminovorans
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
additional information
less than 2% reactivity with N-methyl-iminodiacetate, N-acetamido-iminodiacetate, N-hydroxyethyl-iminodiacetate, EDTA, trimethylamine, triethanolamine, dimethylamine, dimethylglycine, methylamine, sarcosine, anthranilate, betaine, N-methylglutamate, N-methylalanine, N-methylaspartate, N-acetylglutamate, N-methylphenylalanine, N-methylleucine, N-methyltryptophane, glutamate, aspartate, citrate, proline, imidazoleacetate, asparagine, and ethylenediamine-N,N'-diacetate
715329
Aminobacter aminovorans
?
-
-
-
-
additional information
less than 2% reactivity with N-methyl-iminodiacetate, N-acetamido-iminodiacetate, N-hydroxyethyl-iminodiacetate, EDTA, trimethylamine, triethanolamine, dimethylamine, dimethylglycine, methylamine, sarcosine, anthranilate, betaine, N-methylglutamate, N-methylalanine, N-methylaspartate, N-acetylglutamate, N-methylphenylalanine, N-methylleucine, N-methyltryptophane, glutamate, aspartate, citrate, proline, imidazoleacetate, asparagine, and ethylenediamine-N,N'-diacetate
715329
Aminobacter aminovorans ATCC 29600
?
-
-
-
-
nitrilotriacetate + FMNH2 + H+ + O2
specific substrate
715329
Aminobacter aminovorans
iminodiacetate + glyoxylate + FMN + H2O
-
-
-
?
nitrilotriacetate + FMNH2 + H+ + O2
specific substrate
715329
Aminobacter aminovorans ATCC 29600
iminodiacetate + glyoxylate + FMN + H2O
-
-
-
?
Subunits
Subunits
Commentary
Organism
homodimer
2 * 36000, component B of NTA monooxygenase, SDS-PAGE
Aminobacter aminovorans
Temperature Optimum [C]
Temperature Optimum [C]
Temperature Optimum Maximum [C]
Commentary
Organism
25
-
12% activity is left at 2C, and no activity is observed at 40C
Aminobacter aminovorans
pH Optimum
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7.8
-
pH optimum for NTA consumption in cell extracts. At pH 7.2 and 8.4, 25% of the maximum activity is obtained, and no activity is left at pH 6.0 or 9.0
Aminobacter aminovorans
8.5
-
pH optimum for the purified enzyme
Aminobacter aminovorans
pH Range
pH Minimum
pH Maximum
Commentary
Organism
7
9.6
35% activity of the purified enzyme remains at pH 7.0 and 9.6
Aminobacter aminovorans
Cofactor
Cofactor
Commentary
Organism
Structure
FMN
omission of FMN or replacement of FMN by FAD results in a residual activity of 2% when component B is not resaturated with FMN
Aminobacter aminovorans
NADH
NADPH cannot replace NADH
Aminobacter aminovorans
Activating Compound (protein specific)
Activating Compound
Commentary
Organism
Structure
component A of NTA monooxygenase
the ability of component B of NTA monooxygenase to catalyze the oxidation of nitrilotriacetate to iminodiacetate and glyoxylate is completely dependent on the presence of component A
Aminobacter aminovorans
FMN
NTA-Mo activity increases by a factor of 4 when 0.003 mM FMN is included in the assay mixture, there is 0.4 mol of FMN per mol of purified component B
Aminobacter aminovorans
additional information
the activity is increased neither by additions of 20% (w/v) polyethylene glycol or sorbitol
Aminobacter aminovorans
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
FMN
omission of FMN or replacement of FMN by FAD results in a residual activity of 2% when component B is not resaturated with FMN
Aminobacter aminovorans
NADH
NADPH cannot replace NADH
Aminobacter aminovorans
Inhibitors (protein specific)
Inhibitors
Commentary
Organism
Structure
5,5'-dithiobis(2-nitrobenzoate)
67% residual activity at 1 mM
Aminobacter aminovorans
Ca2+
5% residual activity at 10 mM
Aminobacter aminovorans
EDTA
3% residual activity at 10 mM
Aminobacter aminovorans
FAD
24% residual activity at 0.1 mM
Aminobacter aminovorans
NaCl
54% residual activity at 250 mM
Aminobacter aminovorans
p-hydroxymercuribenzoate
12% residual activity at 1 mM
Aminobacter aminovorans
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.0013
-
FMN
in 40 mM TAPS buffer at pH 8.5 and 25C
Aminobacter aminovorans
0.5
-
nitrilotriacetate
in 40 mM TAPS buffer at pH 8.5 and 25C
Aminobacter aminovorans
Metals/Ions (protein specific)
Metals/Ions
Commentary
Organism
Structure
Co2+
may replace Mg2+, enzyme activity is reduced to 80% of that with MgCl2
Aminobacter aminovorans
Mg2+
required, 2 mM used in assay conditions
Aminobacter aminovorans
Mn2+
Mn2+ ions are able to replace Mg2+ but lead to a higher uncoupled NADH oxidation and enzyme activity is reduced to 70% of that with MgCl2
Aminobacter aminovorans
additional information
no nitrilotriacetate consumption is observed with Ca2+,Fe2+, Fe3+, Zn2+, Cu2+, or Ni2+; the enzyme contains less than 0.15 atom of Fe per mol of protein, indicating that neither Fe-sulfur clusters nor cytochromes are present
Aminobacter aminovorans
Molecular Weight [Da] (protein specific)
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
36000
-
2 * 36000, component B of NTA monooxygenase, SDS-PAGE
Aminobacter aminovorans
Purification (Commentary) (protein specific)
Commentary
Organism
ammonium sulfate precipitation, phenyl Sepharose column chromatography, TMAE-Fractogel column chromatography, and phenyl Superose column chromatography
Aminobacter aminovorans
Storage Stability (protein specific)
Storage Stability
Organism
-70C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol, at pH 7.8, several months, without any significant loss of activity
Aminobacter aminovorans
4C, crude extract, 50 mM Tris-HCl, 200 h, 50% loss of activity
Aminobacter aminovorans
4C, purified component B of NTA monooxygenase in 20 mM HEPES buffer at pH 7.8, 7 days, 70% loss of activity
Aminobacter aminovorans
4C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol and ammonium sulfate to 5% saturation, at pH 7.8, 7 days, 5% loss of activity
Aminobacter aminovorans
4C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol, at pH 7.8, 7 days, 20% loss of activity
Aminobacter aminovorans
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
additional information
less than 2% reactivity with N-methyl-iminodiacetate, N-acetamido-iminodiacetate, N-hydroxyethyl-iminodiacetate, EDTA, trimethylamine, triethanolamine, dimethylamine, dimethylglycine, methylamine, sarcosine, anthranilate, betaine, N-methylglutamate, N-methylalanine, N-methylaspartate, N-acetylglutamate, N-methylphenylalanine, N-methylleucine, N-methyltryptophane, glutamate, aspartate, citrate, proline, imidazoleacetate, asparagine, and ethylenediamine-N,N'-diacetate
715329
Aminobacter aminovorans
?
-
-
-
-
additional information
less than 2% reactivity with N-methyl-iminodiacetate, N-acetamido-iminodiacetate, N-hydroxyethyl-iminodiacetate, EDTA, trimethylamine, triethanolamine, dimethylamine, dimethylglycine, methylamine, sarcosine, anthranilate, betaine, N-methylglutamate, N-methylalanine, N-methylaspartate, N-acetylglutamate, N-methylphenylalanine, N-methylleucine, N-methyltryptophane, glutamate, aspartate, citrate, proline, imidazoleacetate, asparagine, and ethylenediamine-N,N'-diacetate
715329
Aminobacter aminovorans ATCC 29600
?
-
-
-
-
nitrilotriacetate + FMNH2 + H+ + O2
specific substrate
715329
Aminobacter aminovorans
iminodiacetate + glyoxylate + FMN + H2O
-
-
-
?
nitrilotriacetate + FMNH2 + H+ + O2
specific substrate
715329
Aminobacter aminovorans ATCC 29600
iminodiacetate + glyoxylate + FMN + H2O
-
-
-
?
Subunits (protein specific)
Subunits
Commentary
Organism
homodimer
2 * 36000, component B of NTA monooxygenase, SDS-PAGE
Aminobacter aminovorans
Temperature Optimum [C] (protein specific)
Temperature Optimum [C]
Temperature Optimum Maximum [C]
Commentary
Organism
25
-
12% activity is left at 2C, and no activity is observed at 40C
Aminobacter aminovorans
pH Optimum (protein specific)
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7.8
-
pH optimum for NTA consumption in cell extracts. At pH 7.2 and 8.4, 25% of the maximum activity is obtained, and no activity is left at pH 6.0 or 9.0
Aminobacter aminovorans
8.5
-
pH optimum for the purified enzyme
Aminobacter aminovorans
pH Range (protein specific)
pH Minimum
pH Maximum
Commentary
Organism
7
9.6
35% activity of the purified enzyme remains at pH 7.0 and 9.6
Aminobacter aminovorans
Other publictions for EC 1.14.14.10
No.
1st author
Pub Med
title
organims
journal
volume
pages
year
Activating Compound
Application
Cloned(Commentary)
Crystallization (Commentary)
Engineering
General Stability
Inhibitors
KM Value [mM]
Localization
Metals/Ions
Molecular Weight [Da]
Natural Substrates/ Products (Substrates)
Organic Solvent Stability
Organism
Oxidation Stability
Posttranslational Modification
Purification (Commentary)
Reaction
Renatured (Commentary)
Source Tissue
Specific Activity [micromol/min/mg]
Storage Stability
Substrates and Products (Substrate)
Subunits
Temperature Optimum [C]
Temperature Range [C]
Temperature Stability [C]
Turnover Number [1/s]
pH Optimum
pH Range
pH Stability
Cofactor
Ki Value [mM]
pI Value
IC50 Value
Activating Compound (protein specific)
Application (protein specific)
Cloned(Commentary) (protein specific)
Cofactor (protein specific)
Crystallization (Commentary) (protein specific)
Engineering (protein specific)
General Stability (protein specific)
IC50 Value (protein specific)
Inhibitors (protein specific)
Ki Value [mM] (protein specific)
KM Value [mM] (protein specific)
Localization (protein specific)
Metals/Ions (protein specific)
Molecular Weight [Da] (protein specific)
Natural Substrates/ Products (Substrates) (protein specific)
Organic Solvent Stability (protein specific)
Oxidation Stability (protein specific)
Posttranslational Modification (protein specific)
Purification (Commentary) (protein specific)
Renatured (Commentary) (protein specific)
Source Tissue (protein specific)
Specific Activity [micromol/min/mg] (protein specific)
Storage Stability (protein specific)
Substrates and Products (Substrate) (protein specific)
Subunits (protein specific)
Temperature Optimum [C] (protein specific)
Temperature Range [C] (protein specific)
Temperature Stability [C] (protein specific)
Turnover Number [1/s] (protein specific)
pH Optimum (protein specific)
pH Range (protein specific)
pH Stability (protein specific)
pI Value (protein specific)
Expression
General Information
General Information (protein specific)
Expression (protein specific)
KCat/KM [mM/s]
KCat/KM [mM/s] (protein specific)
728259
Van Hamme
Genomic and proteomic characte ...
Gordonia sp., Gordonia sp. NB4-1Y
Microbiology
159
1618-1628
2013
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4
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717038
Zhang
Structure of nitrilotriacetate ...
Mycolicibacterium thermoresistibile
Acta Crystallogr. Sect. F
67
1100-1105
2011
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-
1
1
-
-
-
-
-
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6
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1
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1
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1
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1
1
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1
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1
1
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717028
Kim
Crystallization and initial cr ...
Corynebacterium glutamicum
Acta Crystallogr. Sect. F
62
1141-1143
2006
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-
1
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1
2
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2
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1
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1
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717178
Ternan
Iminodiacetate and nitrilotria ...
Kluyveromyces marxianus, Kluyveromyces marxianus IMB3
Biochem. Biophys. Res. Commun.
290
802-805
2002
-
-
-
-
-
1
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1
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4
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1
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2
2
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1
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1
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1
-
2
2
-
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-
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-
-
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717720
Payne
Purification and characterizat ...
Chelativorans sp. BNC1
J. Bacteriol.
180
3823-3827
1998
-
-
-
-
-
-
-
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1
1
-
-
3
-
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1
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2
1
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1
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1
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1
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2
1
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717717
Xu
Cloning, sequencing, and analy ...
Aminobacter aminovorans, Aminobacter aminovorans ATCC 29600
J. Bacteriol.
179
1112-1116
1997
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-
1
-
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1
1
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5
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4
1
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1
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1
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1
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4
1
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717116
Bally
Dynamics of substrate consumpt ...
Aminobacter aminovorans, Aminobacter aminovorans ATCC 29600
Appl. Environ. Microbiol.
62
133-140
1996
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6
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2
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2
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2
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717545
Xun
-
Degradation of metal - nitrilo ...
Aminobacter aminovorans, Aminobacter aminovorans ATCC 29600
Environ. Sci. Technol.
30
1752-1755
1996
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2
7
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7
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5
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1
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1
4
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1
4
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717715
Knobel
Cloning and characterization o ...
Aminobacter aminovorans, Aminobacter aminovorans ATCC 29600
J. Bacteriol.
178
6123-6132
1996
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1
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1
1
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6
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1
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2
1
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1
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1
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1
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1
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1
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1
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718086
Bally
Growth and regulation of enzym ...
Aminobacter aminovorans, Aminobacter aminovorans ATCC 29600
Microbiology
140
1927-1936
1994
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1
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6
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2
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1
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1
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-
715329
Uetz
Purification and characterizat ...
Aminobacter aminovorans, Aminobacter aminovorans ATCC 29600
J. Bacteriol.
174
1179-1188
1992
3
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6
2
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1
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5
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1
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4
1
1
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2
1
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1
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1
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