BRENDA - Enzyme Database show
show all sequences of 1.20.1.1

Relaxing the nicotinamide cofactor specificity of phosphite dehydrogenase by rational design

Woodyer, R.; van der Donk, W.A.; Zhao, H.; Biochemistry 42, 11604-11614 (2003)

Data extracted from this reference:

Application
Application
Commentary
Organism
synthesis
production of deuterium- or tritium-labeled substances, mutant enzymes could be applied as NADPH regeneration systems
Pseudomonas stutzeri
Cloned(Commentary)
Commentary
Organism
wild type and mutant proteins expressed in Escherichia coli BL21(DE3) as His-tag fusion protein
Pseudomonas stutzeri
Engineering
Amino acid exchange
Commentary
Organism
A176R
strongly decreased Km for NADP+
Pseudomonas stutzeri
E175A
strongly decreased Km for NADP+
Pseudomonas stutzeri
E175A/A176R
double mutant, 3.6-fold higher efficiency with NAD+, 1000-fold higher efficiency with NADP+, 3-fold favor for NADP+ over NAD+ as cofactor
Pseudomonas stutzeri
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.004
-
NADP+
25C, pH 7.25, E175A/A176R mutant enzyme
Pseudomonas stutzeri
0.016
-
NAD+
25C, pH 7.25, E175A mutant enzyme
Pseudomonas stutzeri
0.02
-
NAD+
25C, pH 7.25, E175A/A176R mutant enzyme
Pseudomonas stutzeri
0.053
-
NAD+
25C, pH 7.25, wild type enzyme
Pseudomonas stutzeri
0.06
-
NAD+
25C, pH 7.25, A176R mutant enzyme
Pseudomonas stutzeri
0.077
-
NADP+
25C, pH 7.25, A176R mutant enzyme
Pseudomonas stutzeri
0.144
-
NADP+
25C, pH 7.25, E175A mutant enzyme
Pseudomonas stutzeri
2.51
-
NADP+
25C, pH 7.25, wild type enzyme
Pseudomonas stutzeri
Molecular Weight [Da]
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
38500
-
x * 38500, SDS-PAGE, His-tag fusion protein
Pseudomonas stutzeri
Natural Substrates/ Products (Substrates)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
phosphonate + H2O + NAD+
Pseudomonas stutzeri
NADH regeneration system
phosphate + NADH
-
-
ir
Organism
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
Pseudomonas stutzeri
-
-
-
Purification (Commentary)
Commentary
Organism
recombinant enzymes using His-tag
Pseudomonas stutzeri
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
phosphonate + H2O + NAD+
-
657956
Pseudomonas stutzeri
phosphate + NADH
-
-
-
ir
phosphonate + H2O + NAD+
NADH regeneration system
657956
Pseudomonas stutzeri
phosphate + NADH
-
-
-
ir
phosphonate + H2O + NADP+
100-fold lower activity than with NAD+
657956
Pseudomonas stutzeri
phosphate + NADPH
-
-
-
ir
Subunits
Subunits
Commentary
Organism
?
x * 38500, SDS-PAGE, His-tag fusion protein
Pseudomonas stutzeri
Temperature Stability [C]
Temperature Stability Minimum [C]
Temperature Stability Maximum [C]
Commentary
Organism
40.5
-
rapid inactivation, half life less than 10 min, 1 mM NAD+ partially protects against thermal inactivation, E175A/A176R double mutant is readily protected against inactivation by addition of 1 mM NADP+ for at least 15 min
Pseudomonas stutzeri
Turnover Number [1/s]
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
1.41
-
NADP+
25C, pH 7.25, wild type enzyme
Pseudomonas stutzeri
1.9
-
NADP+
25C, pH 7.25, E175A/A176R mutant enzyme
Pseudomonas stutzeri
2.18
-
NADP+
25C, pH 7.25, A176R mutant enzyme; 25C, pH 7.25, E175A mutant enzyme
Pseudomonas stutzeri
2.93
-
NAD+
25C, pH 7.25, wild type enzyme
Pseudomonas stutzeri
3.5
-
NAD+
25C, pH 7.25, E175A mutant enzyme
Pseudomonas stutzeri
3.94
-
NAD+
25C, pH 7.25, E175A/A176R mutant enzyme
Pseudomonas stutzeri
4.28
-
NAD+
25C, pH 7.25, A176R mutant enzyme
Pseudomonas stutzeri
Cofactor
Cofactor
Commentary
Organism
Structure
NAD+
-
Pseudomonas stutzeri
NADP+
100-fold lower activity than with NAD+
Pseudomonas stutzeri
pI Value
Organism
Commentary
pI Value Maximum
pI Value
Pseudomonas stutzeri
isoelectric focusing, wild type enzyme
-
5.8
Pseudomonas stutzeri
isoelectric focusing, A176R mutant enzyme; isoelectric focusing, E175A mutant enzyme
-
6.2
Pseudomonas stutzeri
isoelectric focusing, E175A/A176R mutant enzyme
-
6.6
Application (protein specific)
Application
Commentary
Organism
synthesis
production of deuterium- or tritium-labeled substances, mutant enzymes could be applied as NADPH regeneration systems
Pseudomonas stutzeri
Cloned(Commentary) (protein specific)
Commentary
Organism
wild type and mutant proteins expressed in Escherichia coli BL21(DE3) as His-tag fusion protein
Pseudomonas stutzeri
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
NAD+
-
Pseudomonas stutzeri
NADP+
100-fold lower activity than with NAD+
Pseudomonas stutzeri
Engineering (protein specific)
Amino acid exchange
Commentary
Organism
A176R
strongly decreased Km for NADP+
Pseudomonas stutzeri
E175A
strongly decreased Km for NADP+
Pseudomonas stutzeri
E175A/A176R
double mutant, 3.6-fold higher efficiency with NAD+, 1000-fold higher efficiency with NADP+, 3-fold favor for NADP+ over NAD+ as cofactor
Pseudomonas stutzeri
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.004
-
NADP+
25C, pH 7.25, E175A/A176R mutant enzyme
Pseudomonas stutzeri
0.016
-
NAD+
25C, pH 7.25, E175A mutant enzyme
Pseudomonas stutzeri
0.02
-
NAD+
25C, pH 7.25, E175A/A176R mutant enzyme
Pseudomonas stutzeri
0.053
-
NAD+
25C, pH 7.25, wild type enzyme
Pseudomonas stutzeri
0.06
-
NAD+
25C, pH 7.25, A176R mutant enzyme
Pseudomonas stutzeri
0.077
-
NADP+
25C, pH 7.25, A176R mutant enzyme
Pseudomonas stutzeri
0.144
-
NADP+
25C, pH 7.25, E175A mutant enzyme
Pseudomonas stutzeri
2.51
-
NADP+
25C, pH 7.25, wild type enzyme
Pseudomonas stutzeri
Molecular Weight [Da] (protein specific)
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
38500
-
x * 38500, SDS-PAGE, His-tag fusion protein
Pseudomonas stutzeri
Natural Substrates/ Products (Substrates) (protein specific)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
phosphonate + H2O + NAD+
Pseudomonas stutzeri
NADH regeneration system
phosphate + NADH
-
-
ir
Purification (Commentary) (protein specific)
Commentary
Organism
recombinant enzymes using His-tag
Pseudomonas stutzeri
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
phosphonate + H2O + NAD+
-
657956
Pseudomonas stutzeri
phosphate + NADH
-
-
-
ir
phosphonate + H2O + NAD+
NADH regeneration system
657956
Pseudomonas stutzeri
phosphate + NADH
-
-
-
ir
phosphonate + H2O + NADP+
100-fold lower activity than with NAD+
657956
Pseudomonas stutzeri
phosphate + NADPH
-
-
-
ir
Subunits (protein specific)
Subunits
Commentary
Organism
?
x * 38500, SDS-PAGE, His-tag fusion protein
Pseudomonas stutzeri
Temperature Stability [C] (protein specific)
Temperature Stability Minimum [C]
Temperature Stability Maximum [C]
Commentary
Organism
40.5
-
rapid inactivation, half life less than 10 min, 1 mM NAD+ partially protects against thermal inactivation, E175A/A176R double mutant is readily protected against inactivation by addition of 1 mM NADP+ for at least 15 min
Pseudomonas stutzeri
Turnover Number [1/s] (protein specific)
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
1.41
-
NADP+
25C, pH 7.25, wild type enzyme
Pseudomonas stutzeri
1.9
-
NADP+
25C, pH 7.25, E175A/A176R mutant enzyme
Pseudomonas stutzeri
2.18
-
NADP+
25C, pH 7.25, A176R mutant enzyme; 25C, pH 7.25, E175A mutant enzyme
Pseudomonas stutzeri
2.93
-
NAD+
25C, pH 7.25, wild type enzyme
Pseudomonas stutzeri
3.5
-
NAD+
25C, pH 7.25, E175A mutant enzyme
Pseudomonas stutzeri
3.94
-
NAD+
25C, pH 7.25, E175A/A176R mutant enzyme
Pseudomonas stutzeri
4.28
-
NAD+
25C, pH 7.25, A176R mutant enzyme
Pseudomonas stutzeri
pI Value (protein specific)
Organism
Commentary
pI Value Maximum
pI Value
Pseudomonas stutzeri
isoelectric focusing, wild type enzyme
-
5.8
Pseudomonas stutzeri
isoelectric focusing, A176R mutant enzyme; isoelectric focusing, E175A mutant enzyme
-
6.2
Pseudomonas stutzeri
isoelectric focusing, E175A/A176R mutant enzyme
-
6.6
Other publictions for EC 1.20.1.1
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)
744126
Beyer
P450BM3 fused to phosphite de ...
Pseudomonas stutzeri, Pseudomonas stutzeri WM88
Appl. Microbiol. Biotechnol.
101
2319-2331
2017
-
-
1
-
-
-
-
1
-
-
-
2
-
5
-
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1
-
-
-
-
-
2
-
-
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-
1
-
-
-
1
-
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-
1
1
-
-
-
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1
-
-
-
2
-
-
-
1
-
-
-
-
2
-
-
-
-
1
-
-
-
-
-
-
-
-
1
1
744691
Ranaghan
-
A catalytic role for methioni ...
Pseudomonas stutzeri, Pseudomonas stutzeri WM88
Chem. Sci.
5
2191-2199
2014
-
-
-
-
3
-
1
6
-
-
-
2
-
3
-
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2
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3
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1
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1
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3
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1
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6
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2
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-
-
2
-
-
-
-
3
-
-
-
-
-
-
-
-
3
3
745418
Kanda
Application of a phosphite de ...
Ralstonia sp.
J. Biotechnol.
182-183
68-73
2014
-
-
1
-
-
-
-
-
-
-
-
1
-
4
-
-
-
-
-
-
-
-
1
-
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-
-
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1
-
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-
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1
1
-
-
-
-
-
-
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-
1
-
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-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
746256
Hung
Chemical rescue and inhibitio ...
Pseudomonas stutzeri, Pseudomonas stutzeri WM88
PLoS ONE
9
e87134
2014
-
-
-
1
2
-
1
3
-
-
-
2
-
4
-
-
1
-
-
-
-
-
2
-
-
-
-
-
-
-
-
1
2
-
-
-
-
-
1
1
2
-
-
1
2
3
-
-
-
2
-
-
-
1
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
726694
Liu
Cloning, expression, and chara ...
Pseudomonas sp., Pseudomonas sp. K
Appl. Biochem. Biotechnol.
166
1301-1313
2012
-
-
1
1
-
-
4
2
-
3
2
-
-
6
-
-
1
-
-
-
1
-
2
1
1
2
3
-
1
2
1
-
-
-
-
-
-
1
-
1
-
-
-
4
-
2
-
3
2
-
-
-
-
1
-
-
1
-
2
1
1
2
3
-
1
2
1
-
-
-
-
-
-
-
726981
Hung
Investigation of the role of A ...
Cupriavidus metallidurans, Cupriavidus metallidurans CH34, Methylorubrum extorquens, Methylorubrum extorquens ATCC 14718 / DSM 1338 / JCM 2805 / NCIMB 9133 / AM1, Nostoc punctiforme, Pseudomonas stutzeri
Biochemistry
51
4254-4262
2012
-
-
4
-
28
-
4
22
-
-
-
-
-
25
-
-
-
-
-
-
-
-
6
-
-
-
-
11
-
-
-
-
3
-
-
-
-
4
-
-
28
-
-
4
3
22
-
-
-
-
-
-
-
-
-
-
-
-
6
-
-
-
-
11
-
-
-
-
-
-
-
-
7
7
726982
Zou
Crystal structures of phosphit ...
Pseudomonas stutzeri
Biochemistry
51
4263-4270
2012
-
-
1
1
1
-
1
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
1
1
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
728004
Hirota
Isolation and characterization ...
Pseudomonas stutzeri, Pseudomonas stutzeri WM88, Ralstonia sp., Ralstonia sp. 4506
J. Biosci. Bioeng.
113
445-450
2012
-
-
2
-
-
-
7
8
1
-
-
-
-
11
-
-
-
-
-
-
-
-
4
-
2
-
2
-
-
-
-
-
-
-
-
-
-
2
-
-
-
-
-
7
-
8
1
-
-
-
-
-
-
-
-
-
-
-
4
-
2
-
2
-
-
-
-
-
-
-
-
-
-
-
691424
McLachlan
Further improvement of phosphi ...
Pseudomonas stutzeri
Biotechnol. Bioeng.
99
268-274
2008
-
1
1
-
33
-
-
10
-
-
-
-
-
1
-
-
1
-
-
-
-
1
1
-
-
-
-
4
-
-
-
1
-
-
-
-
1
1
1
-
33
-
-
-
-
10
-
-
-
-
-
-
-
1
-
-
-
1
1
-
-
-
-
4
-
-
-
-
-
-
-
-
-
-
672712
Johannes
Efficient regeneration of NADP ...
Pseudomonas stutzeri
Biotechnol. Bioeng.
96
18-26
2007
-
1
-
-
1
-
-
-
-
-
-
-
-
4
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
1
-
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-
-
-
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-
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-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
690884
Fogle
Pre-steady-state studies of ph ...
Pseudomonas stutzeri, Pseudomonas stutzeri WW88
Biochemistry
46
13101-13108
2007
-
1
1
-
6
-
1
8
-
-
-
-
-
2
-
-
1
-
-
-
-
-
2
-
-
-
-
9
-
-
-
1
-
-
-
-
1
1
1
-
6
-
-
1
-
8
-
-
-
-
-
-
-
1
-
-
-
-
2
-
-
-
-
9
-
-
-
-
-
-
-
-
-
-
673144
Woodyer
Optimizing a biocatalyst for i ...
Pseudomonas stutzeri
Comb. Chem. High Throughput Screen.
9
237-245
2006
-
1
-
-
7
-
-
25
-
-
-
-
-
2
-
-
-
-
-
-
-
-
2
-
-
-
-
12
-
-
-
-
-
-
-
-
1
-
-
-
7
-
-
-
-
25
-
-
-
-
-
-
-
-
-
-
-
-
2
-
-
-
-
12
-
-
-
-
-
-
-
-
-
-
658120
Woodyer
Site-directed mutagenesis of a ...
Pseudomonas stutzeri, Pseudomonas stutzeri WM 88
Biochemistry
44
4765-4774
2005
-
-
1
-
12
-
2
13
-
-
2
2
-
2
-
-
1
-
-
-
-
-
4
1
-
-
-
7
1
1
-
1
-
-
-
-
-
1
1
-
12
-
-
2
-
13
-
-
2
2
-
-
-
1
-
-
-
-
4
1
-
-
-
7
1
1
-
-
-
-
-
-
-
-
658126
Relyea
Inhibition and pH dependence o ...
Pseudomonas stutzeri
Biochemistry
44
6640-6649
2005
-
-
-
-
-
-
1
-
-
-
-
1
-
1
-
-
-
-
-
-
-
-
3
-
-
-
-
-
1
1
-
1
-
-
-
-
-
-
1
-
-
-
-
1
-
-
-
-
-
1
-
-
-
-
-
-
-
-
3
-
-
-
-
-
1
1
-
-
-
-
-
-
-
-
658259
Relyea
Mechanism and applications of ...
Alcaligenes faecalis, Pseudomonas stutzeri, Pseudomonas stutzeri WM88
Bioorg. Chem.
33
171-189
2005
-
1
-
-
6
-
2
-
-
-
-
3
-
5
-
-
-
1
-
-
-
-
10
-
1
-
-
-
1
-
-
3
-
-
-
-
1
-
3
-
6
-
-
2
-
-
-
-
-
3
-
-
-
-
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-
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10
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1
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1
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671425
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1
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17
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33
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3
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-
-
-
-
-
-
-
1
-
-
-
2
12
-
-
-
-
-
-
-
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1
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-
-
17
-
-
-
-
33
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
2
12
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-
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673526
Woodyer
Mechanistic investigation of a ...
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2005
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1
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1
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-
-
-
-
-
-
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2
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
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-
-
-
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1
-
-
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1
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-
-
-
-
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-
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-
-
-
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-
-
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657956
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Relaxing the nicotinamide cofa ...
Pseudomonas stutzeri
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1
1
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3
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8
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1
1
-
2
-
-
1
-
-
-
-
-
3
1
-
-
1
7
-
-
-
2
-
3
-
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1
1
2
-
3
-
-
-
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8
-
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1
1
-
-
-
1
-
-
-
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3
1
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1
7
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2001
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1
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2
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1
1
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3
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1
-
-
-
1
-
4
1
-
-
-
1
1
1
-
-
-
-
-
-
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1
-
-
-
-
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9
-
2
-
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1
1
-
-
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1
-
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1
-
4
1
-
-
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1
1
1
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287854
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Phosphite dehydrogenase: An un ...
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1
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1
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4
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2
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1
1
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1
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1
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1
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4
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2
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1
1
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