BRENDA - Enzyme Database show
show all sequences of 1.1.1.B4

Cofactor engineering of Lactobacillus brevis alcohol dehydrogenase by computational design

Machielsen, R.; Looger, L.; Raedts, J.; Dijkhuizen, S.; Hummel, W.; Henneman, H.; Daussmann, T.; van der Oost, J.; Eng. Life Sci. 9, 38-44 (2009)
No PubMed abstract available

Data extracted from this reference:

Cloned(Commentary)
Commentary
Organism
expression in Escherichia coli
Lactobacillus brevis
Engineering
Amino acid exchange
Commentary
Organism
A90S
activity with NADPH as cofactor decreased to about 50% of wild-type, activity with NADH strongly decreased
Lactobacillus brevis
G37D/R38P
activity with NADH is decreased relative to Arg38Pro alone, but is higher than that of the wild-type enzyme
Lactobacillus brevis
M140I
no activity with NADPH as cofactor
Lactobacillus brevis
R38P
no activity with NADPH as cofactor, fourfold increase in activity with NADH
Lactobacillus brevis
V112D
no activity with NADPH as cofactor, strongly decreased activity with NADH
Lactobacillus brevis
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.017
-
NADPH
wild-type, pH 6.5, 30°C
Lactobacillus brevis
0.028
-
NADPH
mutant R38P, pH 5.5, 30°C
Lactobacillus brevis
0.12
-
NADH
mutant R38P, pH 5.5, 30°C
Lactobacillus brevis
0.13
-
NADH
wild-type, pH 6.5, 30°C
Lactobacillus brevis
Organism
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
Lactobacillus brevis
-
-
-
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
acetophenone + NADH + H+
-
711860
Lactobacillus brevis
phenylethanol + NAD+
-
-
-
?
acetophenone + NADPH + H+
-
711860
Lactobacillus brevis
phenylethanol + NADP+
-
-
-
?
Temperature Stability [°C]
Temperature Stability Minimum [°C]
Temperature Stability Maximum [°C]
Commentary
Organism
30
-
wild-type, half-life 160 h, mutant R38P, half-life 220 h
Lactobacillus brevis
42
-
wild-type, half-life 1.25 h, mutant R38P, half-life 3 h
Lactobacillus brevis
pH Optimum
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
5.5
-
wild-type, cofactor NADH. Mutant R38P, both cofactors NADH and NADPH
Lactobacillus brevis
6
6.5
wild-type, cofactor NADPH
Lactobacillus brevis
Cofactor
Cofactor
Commentary
Organism
Structure
NADPH
clear preference for NADPH as cofactor. The phosphate group of NADPH interacts with both the side chains and main-chain amides of Arg38 and Thr15, stabilizing this cofactor significantly better than NADH
Lactobacillus brevis
Cloned(Commentary) (protein specific)
Commentary
Organism
expression in Escherichia coli
Lactobacillus brevis
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
NADPH
clear preference for NADPH as cofactor. The phosphate group of NADPH interacts with both the side chains and main-chain amides of Arg38 and Thr15, stabilizing this cofactor significantly better than NADH
Lactobacillus brevis
Engineering (protein specific)
Amino acid exchange
Commentary
Organism
A90S
activity with NADPH as cofactor decreased to about 50% of wild-type, activity with NADH strongly decreased
Lactobacillus brevis
G37D/R38P
activity with NADH is decreased relative to Arg38Pro alone, but is higher than that of the wild-type enzyme
Lactobacillus brevis
M140I
no activity with NADPH as cofactor
Lactobacillus brevis
R38P
no activity with NADPH as cofactor, fourfold increase in activity with NADH
Lactobacillus brevis
V112D
no activity with NADPH as cofactor, strongly decreased activity with NADH
Lactobacillus brevis
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.017
-
NADPH
wild-type, pH 6.5, 30°C
Lactobacillus brevis
0.028
-
NADPH
mutant R38P, pH 5.5, 30°C
Lactobacillus brevis
0.12
-
NADH
mutant R38P, pH 5.5, 30°C
Lactobacillus brevis
0.13
-
NADH
wild-type, pH 6.5, 30°C
Lactobacillus brevis
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
acetophenone + NADH + H+
-
711860
Lactobacillus brevis
phenylethanol + NAD+
-
-
-
?
acetophenone + NADPH + H+
-
711860
Lactobacillus brevis
phenylethanol + NADP+
-
-
-
?
Temperature Stability [°C] (protein specific)
Temperature Stability Minimum [°C]
Temperature Stability Maximum [°C]
Commentary
Organism
30
-
wild-type, half-life 160 h, mutant R38P, half-life 220 h
Lactobacillus brevis
42
-
wild-type, half-life 1.25 h, mutant R38P, half-life 3 h
Lactobacillus brevis
pH Optimum (protein specific)
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
5.5
-
wild-type, cofactor NADH. Mutant R38P, both cofactors NADH and NADPH
Lactobacillus brevis
6
6.5
wild-type, cofactor NADPH
Lactobacillus brevis
KCat/KM [mM/s]
kcat/KM Value [1/mMs-1]
kcat/KM Value Maximum [1/mMs-1]
Substrate
Commentary
Organism
Structure
75
-
NADH
wild-type, pH 6.5, 30°C
Lactobacillus brevis
300
-
NADH
mutant R38P, pH 5.5, 30°C
Lactobacillus brevis
5100
-
NADPH
mutant R38P, pH 5.5, 30°C
Lactobacillus brevis
9300
-
NADPH
wild-type, pH 6.5, 30°C
Lactobacillus brevis
KCat/KM [mM/s] (protein specific)
KCat/KM Value [1/mMs-1]
KCat/KM Value Maximum [1/mMs-1]
Substrate
Commentary
Organism
Structure
75
-
NADH
wild-type, pH 6.5, 30°C
Lactobacillus brevis
300
-
NADH
mutant R38P, pH 5.5, 30°C
Lactobacillus brevis
5100
-
NADPH
mutant R38P, pH 5.5, 30°C
Lactobacillus brevis
9300
-
NADPH
wild-type, pH 6.5, 30°C
Lactobacillus brevis
Other publictions for EC 1.1.1.B4
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)
721911
Zhou
Biocatalytic characterization ...
Carboxydothermus hydrogenoformans, Carboxydothermus hydrogenoformans DSM 6008
Biotechnol. Lett.
35
359-365
2013
-
1
1
-
-
-
-
1
-
-
1
-
4
2
-
-
-
-
-
-
-
-
15
1
1
-
-
1
1
-
-
2
-
-
-
-
1
1
2
-
-
-
-
-
-
1
-
-
1
-
4
-
-
-
-
-
-
-
15
1
1
-
-
1
1
-
-
-
-
-
-
-
-
-
723841
Protsko
Crystallization and preliminar ...
Thermoanaerobacter ethanolicus
Acta Crystallogr. Sect. F
66
831-833
2010
-
-
1
1
1
-
-
-
-
-
1
1
-
1
-
-
1
-
-
-
-
-
1
1
-
-
-
-
-
-
-
2
-
-
-
-
-
1
2
1
1
-
-
-
-
-
-
-
1
1
-
-
-
1
-
-
-
-
1
1
-
-
-
-
-
-
-
-
-
1
1
-
-
-
725726
Li
-
Deracemization of aryl seconda ...
Microbacterium oxydans, Microbacterium oxydans ECU2010
J. Mol. Catal. B
64
48-52
2010
-
1
-
-
-
-
-
-
-
-
-
2
-
2
-
-
-
-
-
-
-
-
4
-
-
-
-
-
-
-
-
2
-
-
-
-
1
-
2
-
-
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
4
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
697267
Pal
Activity of yeast alcohol dehy ...
Saccharomyces carlsbergensis, Saccharomyces carlsbergensis Y379-50, Saccharomyces cerevisiae
Chem. Biol. Interact.
178
16-23
2009
-
-
-
-
-
-
-
-
1
-
-
-
-
3
-
-
-
-
-
-
-
-
3
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
2
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
3
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
697839
Pennacchio
Role of Tryptophan 95 in subst ...
Sulfolobus solfataricus
Extremophiles
13
751-761
2009
-
-
1
1
1
-
-
4
-
-
-
-
-
1
-
-
1
-
-
-
-
-
2
-
-
-
-
4
-
-
-
1
-
-
-
-
-
1
1
1
1
-
-
-
-
4
-
-
-
-
-
-
-
1
-
-
-
-
2
-
-
-
-
4
-
-
-
-
-
-
-
-
-
-
711860
Machielsen
-
Cofactor engineering of Lactob ...
Lactobacillus brevis
Eng. Life Sci.
9
38-44
2009
-
-
1
-
5
-
-
4
-
-
-
-
-
1
-
-
-
-
-
-
-
-
2
-
-
-
2
-
2
-
-
1
-
-
-
-
-
1
1
-
5
-
-
-
-
4
-
-
-
-
-
-
-
-
-
-
-
-
2
-
-
-
2
-
2
-
-
-
-
-
-
-
4
4
684583
Pennacchio
Purification and characterizat ...
Thermus thermophilus
Appl. Environ. Microbiol.
74
3949-3958
2008
1
1
1
-
-
-
7
5
-
5
4
-
1
1
-
-
1
-
-
-
-
-
7
1
2
1
4
4
2
2
-
3
-
-
-
1
1
1
3
-
-
-
-
7
-
5
-
5
4
-
1
-
-
1
-
-
-
-
7
1
2
1
4
4
2
2
-
-
-
-
-
-
-
-
688747
Nie
A novel NADH-dependent carbony ...
Candida parapsilosis
Lett. Appl. Microbiol.
44
555-562
2007
-
-
-
-
-
-
9
-
1
1
2
-
-
1
-
-
1
-
-
-
1
1
10
1
1
-
-
-
1
-
-
2
-
-
-
-
-
-
2
-
-
-
-
9
-
-
1
1
2
-
-
-
-
1
-
-
1
1
10
1
1
-
-
-
1
-
-
-
-
-
-
-
-
-
687864
Isobe
Thermostable NAD+-dependent (R ...
Burkholderia sp., Burkholderia sp. AIU 652
J. Biosci. Bioeng.
96
387-393
2003
-
-
-
-
-
-
6
15
-
-
2
-
-
3
-
-
1
-
-
-
1
-
24
1
1
-
1
-
3
-
-
3
-
1
-
-
-
-
3
-
-
-
-
6
-
15
-
-
2
-
-
-
-
1
-
-
1
-
24
1
1
-
1
-
3
-
-
1
-
-
-
-
-
-
688571
Bradshaw
-
A Pseudomonas sp. alcohol dehy ...
Pseudomonas sp., Pseudomonas sp. PED
J. Org. Chem.
57
1526-1532
1992
-
-
-
-
-
-
-
2
-
-
-
-
-
2
-
-
-
1
-
-
1
-
24
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-
-
1
-
24
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-