BRENDA - Enzyme Database
show all sequences of 1.1.99.31

A novel structural basis for membrane association of a protein: construction of a chimeric soluble mutant of (S)-mandelate dehydrogenase from Pseudomonas putida

Mitra, B.; Gerlt, J.A.; Babbitt, P.C.; Koo, C.W.; Kenyon, G.L.; Joseph, D.; Petsko, G.A.; Biochemistry 32, 12959-12967 (1993)

Data extracted from this reference:

Cloned(Commentary)
Cloned (Commentary)
Organism
expression in Escherichia coli
Pseudomonas putida
Engineering
Protein Variants
Commentary
Organism
additional information
chimeric mutant of (S)-mandelate dehydrogenase with membrane anchoring loop replaced by a portion of glycolate oxidase from spinach is soluble and retains partial catalytic activity (about 1%) using (S)-mandelate as substrate. The activities of the soluble mutant enzymes (S)-mandelate dehydrogenase with residues 2-4 deleted and (S)-mandelate dehydrogenase with 17 residues deleted at the carboxy terminus are nearly the same when (S)-phenyllactate is used as substrate
Pseudomonas putida
General Stability
General Stability
Organism
repeated freeze-thaw cycles cause significant loss of activity
Pseudomonas putida
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.158
-
(S)-Mandelate
pH 7.5, (S)-mandelate dehydrogenase with 17 residues deleted at the carboxy terminus
Pseudomonas putida
0.206
-
(S)-Mandelate
pH 7.5, wild-type enzyme
Pseudomonas putida
0.225
-
(S)-Mandelate
pH 7.5, (S)-mandelate dehydrogenase with residues 2-4 deleted
Pseudomonas putida
0.229
-
(S)-Mandelate
pH 7.5, chimeric mutant of (S)-mandelate dehydrogenase with membrane anchoring loop replaced by a portion of glycolate oxidase from spinach
Pseudomonas putida
0.78
-
(S)-phenyllactate
pH 7.5, chimeric mutant of (S)-mandelate dehydrogenase with membrane anchoring loop replaced by a portion of glycolate oxidase from spinach
Pseudomonas putida
2.6
-
(S)-phenyllactate
pH 7.5, wild-type enzyme
Pseudomonas putida
Localization
Localization
Commentary
Organism
GeneOntology No.
Textmining
membrane
associated, wild-type enzyme
Pseudomonas putida
16020
-
Organism
Organism
UniProt
Commentary
Textmining
Pseudomonas putida
-
-
-
Purification (Commentary)
Purification (Commentary)
Organism
wild-type and mutant enzymes
Pseudomonas putida
Storage Stability
Storage Stability
Organism
-70°C, wild-type enzyme can be stored frozen in 20% ethanediol for weeks without loss of activity
Pseudomonas putida
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
Substrate Product ID
(S)-mandelate + 2,6-dichlorophenolindophenol
-
667564
Pseudomonas putida
2-oxo-2-phenylacetate + reduced 2,6-dichlorophenolindophenol
-
-
-
?
(S)-phenyllactate + 2,6-dichlorophenolindophenol
-
667564
Pseudomonas putida
?
-
-
-
?
Synonyms
Synonyms
Commentary
Organism
MDH
-
Pseudomonas putida
Turnover Number [1/s]
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
1.6
-
(S)-Mandelate
pH 7.5, chimeric mutant of (S)-mandelate dehydrogenase with membrane anchoring loop replaced by a portion of glycolate oxidase from spinach
Pseudomonas putida
9
-
(S)-Mandelate
pH 7.5, (S)-mandelate dehydrogenase with residues 2-4 deleted
Pseudomonas putida
24
-
(S)-Mandelate
pH 7.5, (S)-mandelate dehydrogenase with 17 residues deleted at the carboxy terminus
Pseudomonas putida
174
-
(S)-Mandelate
pH 7.5, wild-type enzyme
Pseudomonas putida
Cofactor
Cofactor
Commentary
Organism
Structure
FMN
-
Pseudomonas putida
Cloned(Commentary) (protein specific)
Commentary
Organism
expression in Escherichia coli
Pseudomonas putida
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
FMN
-
Pseudomonas putida
Engineering (protein specific)
Protein Variants
Commentary
Organism
additional information
chimeric mutant of (S)-mandelate dehydrogenase with membrane anchoring loop replaced by a portion of glycolate oxidase from spinach is soluble and retains partial catalytic activity (about 1%) using (S)-mandelate as substrate. The activities of the soluble mutant enzymes (S)-mandelate dehydrogenase with residues 2-4 deleted and (S)-mandelate dehydrogenase with 17 residues deleted at the carboxy terminus are nearly the same when (S)-phenyllactate is used as substrate
Pseudomonas putida
General Stability (protein specific)
General Stability
Organism
repeated freeze-thaw cycles cause significant loss of activity
Pseudomonas putida
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.158
-
(S)-Mandelate
pH 7.5, (S)-mandelate dehydrogenase with 17 residues deleted at the carboxy terminus
Pseudomonas putida
0.206
-
(S)-Mandelate
pH 7.5, wild-type enzyme
Pseudomonas putida
0.225
-
(S)-Mandelate
pH 7.5, (S)-mandelate dehydrogenase with residues 2-4 deleted
Pseudomonas putida
0.229
-
(S)-Mandelate
pH 7.5, chimeric mutant of (S)-mandelate dehydrogenase with membrane anchoring loop replaced by a portion of glycolate oxidase from spinach
Pseudomonas putida
0.78
-
(S)-phenyllactate
pH 7.5, chimeric mutant of (S)-mandelate dehydrogenase with membrane anchoring loop replaced by a portion of glycolate oxidase from spinach
Pseudomonas putida
2.6
-
(S)-phenyllactate
pH 7.5, wild-type enzyme
Pseudomonas putida
Localization (protein specific)
Localization
Commentary
Organism
GeneOntology No.
Textmining
membrane
associated, wild-type enzyme
Pseudomonas putida
16020
-
Purification (Commentary) (protein specific)
Commentary
Organism
wild-type and mutant enzymes
Pseudomonas putida
Storage Stability (protein specific)
Storage Stability
Organism
-70°C, wild-type enzyme can be stored frozen in 20% ethanediol for weeks without loss of activity
Pseudomonas putida
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ID
(S)-mandelate + 2,6-dichlorophenolindophenol
-
667564
Pseudomonas putida
2-oxo-2-phenylacetate + reduced 2,6-dichlorophenolindophenol
-
-
-
?
(S)-phenyllactate + 2,6-dichlorophenolindophenol
-
667564
Pseudomonas putida
?
-
-
-
?
Turnover Number [1/s] (protein specific)
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
1.6
-
(S)-Mandelate
pH 7.5, chimeric mutant of (S)-mandelate dehydrogenase with membrane anchoring loop replaced by a portion of glycolate oxidase from spinach
Pseudomonas putida
9
-
(S)-Mandelate
pH 7.5, (S)-mandelate dehydrogenase with residues 2-4 deleted
Pseudomonas putida
24
-
(S)-Mandelate
pH 7.5, (S)-mandelate dehydrogenase with 17 residues deleted at the carboxy terminus
Pseudomonas putida
174
-
(S)-Mandelate
pH 7.5, wild-type enzyme
Pseudomonas putida
Other publictions for EC 1.1.99.31
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
Synonyms
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)
740174
Chen
Stereoselective biotransformat ...
Escherichia coli
Bioresour. Bioproc.
4
2-2
2017
-
-
-
-
-
-
-
1
-
-
-
1
-
1
-
-
1
-
-
-
-
-
1
-
1
1
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
1
-
-
-
1
-
-
-
-
1
-
1
-
-
-
1
-
-
-
-
-
-
-
-
-
740119
Mazurenko
Immobilization of membrane-bou ...
Pseudomonas putida, Pseudomonas putida DSM 291
Bioelectrochemistry
104
65-70
2015
-
-
-
-
-
-
-
-
-
-
-
2
-
4
-
-
-
-
-
-
-
-
2
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
725744
Wang
-
A bi-enzymatic system for effi ...
Pseudomonas aeruginosa
J. Mol. Catal. B
94
47-50
2013
-
-
1
-
-
-
-
2
-
-
-
1
-
1
-
-
1
-
-
-
2
-
1
-
1
1
1
1
-
1
-
-
1
-
-
-
-
-
1
1
-
-
-
-
-
-
2
-
-
-
1
-
-
-
1
-
-
2
-
1
-
1
1
1
-
1
-
-
-
-
-
-
-
-
-
695329
Sukumar
Structures of the G81A mutant ...
Pseudomonas putida
Acta Crystallogr. Sect. D
65
543-552
2009
-
-
1
1
2
-
-
12
-
-
-
-
-
4
-
-
1
-
-
-
-
-
4
-
1
-
-
-
12
-
-
-
1
-
-
-
-
-
1
1
1
2
-
-
-
-
12
-
-
-
-
-
-
-
1
-
-
-
-
4
-
-
-
-
12
-
-
-
-
-
-
-
-
-
-
667577
Dewanti
Role of glycine 81 in (S)-mand ...
Pseudomonas putida
Biochemistry
43
10692-10700
2004
-
-
-
-
5
-
1
36
-
-
-
-
-
4
-
-
1
-
-
-
-
-
13
-
-
-
-
-
36
-
-
-
1
-
-
-
-
-
-
1
-
5
-
-
1
-
36
-
-
-
-
-
-
-
1
-
-
-
-
13
-
-
-
-
36
-
-
-
-
-
-
-
-
-
-
669259
Sukumar
High resolution structures of ...
Pseudomonas putida
J. Biol. Chem.
279
3749-3757
2004
-
-
-
1
-
-
-
-
-
-
-
-
-
3
-
-
-
-
-
-
-
-
1
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
678128
Dewanti
Esters of mandelic acid as sub ...
Pseudomonas putida
Biochemistry
43
1883-1890
2004
-
-
-
-
2
-
4
9
-
-
-
-
-
5
-
-
1
-
-
-
-
-
4
-
1
-
-
-
9
-
-
-
1
2
-
-
-
-
-
1
-
2
-
-
4
2
9
-
-
-
-
-
-
-
1
-
-
-
-
4
-
-
-
-
9
-
-
-
-
-
-
-
-
-
-
667575
Dewanti
A transient intermediate in th ...
Pseudomonas putida
Biochemistry
42
12893-12901
2003
-
-
-
-
-
-
-
2
-
-
-
-
-
5
-
-
1
-
-
-
-
-
1
-
-
-
-
-
2
-
-
-
1
-
-
-
-
-
-
1
-
-
-
-
-
-
2
-
-
-
-
-
-
-
1
-
-
-
-
1
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
667572
Xu
Arginine 165/arginine 277 pair ...
Pseudomonas putida
Biochemistry
41
12313-12319
2002
-
-
-
-
10
-
3
9
-
-
-
-
-
5
-
-
1
-
-
-
-
1
1
-
-
-
-
-
9
-
-
-
1
13
-
-
-
-
-
1
-
10
-
-
3
13
9
-
-
-
-
-
-
-
1
-
-
-
1
1
-
-
-
-
9
-
-
-
-
-
-
-
-
-
-
667570
Sukumar
Structure of an active soluble ...
Pseudomonas putida
Biochemistry
40
9870-9878
2001
-
-
-
1
-
-
-
-
-
-
-
-
-
3
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
667567
Lehoux
Role of arginine 277 in (S)-ma ...
Pseudomonas putida
Biochemistry
39
10055-10065
2000
-
-
-
-
4
-
-
7
-
-
-
-
-
4
-
-
1
-
-
-
-
-
2
-
-
-
-
-
7
-
-
-
-
-
-
-
-
-
-
-
-
4
-
-
-
-
7
-
-
-
-
-
-
-
1
-
-
-
-
2
-
-
-
-
7
-
-
-
-
-
-
-
-
-
-
667565
Xu
A highly active, soluble mutan ...
Pseudomonas putida
Biochemistry
38
12367-12376
1999
-
-
-
-
1
-
-
14
1
-
-
-
-
6
-
-
-
-
-
-
-
-
14
-
1
-
-
-
15
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
14
1
-
-
-
-
-
-
-
-
-
-
-
14
-
-
-
-
15
-
-
-
-
-
-
-
-
-
-
667566
Lehoux
(S)-Mandelate dehydrogenase fr ...
Pseudomonas putida
Biochemistry
38
9948-9955
1999
-
-
-
-
3
-
-
1
-
-
-
-
-
5
-
-
1
-
-
-
-
-
2
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
-
-
3
-
-
-
-
1
-
-
-
-
-
-
-
1
-
-
-
-
2
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
678122
Lehoux
(S)-Mandelate dehydrogenase fr ...
Pseudomonas putida
Biochemistry
38
5836-5848
1999
-
-
1
-
-
-
5
11
-
-
-
-
-
4
-
-
1
-
-
-
-
-
14
-
1
-
-
-
-
-
-
-
1
1
-
-
-
-
1
1
-
-
-
-
5
1
11
-
-
-
-
-
-
-
1
-
-
-
-
14
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
667458
Illias
L-Mandelate dehydrogenase from ...
Rhodotorula graminis
Biochem. J.
333
107-115
1998
-
-
1
-
-
-
-
6
2
-
-
-
-
8
-
-
1
-
-
-
-
-
3
-
1
-
-
-
8
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
6
2
-
-
-
-
-
-
1
-
-
-
-
3
-
-
-
-
8
-
-
-
-
-
-
-
-
-
-
667457
Smekal
Substrate analogues as probes ...
Rhodotorula graminis
Biochem. J.
297
647-652
1994
-
-
-
-
-
-
-
11
-
-
-
-
-
3
-
-
-
-
-
-
-
-
11
-
-
-
-
-
12
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
11
-
-
-
-
-
-
-
-
-
-
-
-
11
-
-
-
-
12
-
-
-
-
-
-
-
-
-
-
667455
Smekal
L-mandelate dehydrogenase from ...
Rhodotorula graminis
Biochem. J.
290
103-107
1993
-
-
-
-
-
-
4
3
-
-
-
-
-
7
-
-
-
-
-
-
-
-
3
-
2
-
-
-
3
-
-
-
1
4
-
-
-
-
-
1
-
-
-
-
4
4
3
-
-
-
-
-
-
-
-
-
-
-
-
3
-
-
-
-
3
-
-
-
-
-
-
-
-
-
-
667564
Mitra
A novel structural basis for m ...
Pseudomonas putida
Biochemistry
32
12959-12967
1993
-
-
1
-
1
1
-
6
2
-
-
-
-
6
-
-
1
-
-
-
-
1
2
-
1
-
-
-
4
-
-
-
1
-
-
-
-
-
1
1
-
1
1
-
-
-
6
2
-
-
-
-
-
-
1
-
-
-
1
2
-
-
-
-
4
-
-
-
-
-
-
-
-
-
-
669678
Fewson
Comparison of mandelate dehydr ...
Acinetobacter calcoaceticus
J. Gen. Microbiol.
134
967-974
1988
-
-
-
-
-
-
1
-
-
-
-
-
-
4
-
-
-
-
-
-
-
-
1
-
2
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
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-
1
-
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