BRENDA - Enzyme Database
show all sequences of 1.14.11.55

Molecular dynamics simulations and structure-guided mutagenesis provide insight into the architecture of the catalytic core of the ectoine hydroxylase

Widderich, N.; Pittelkow, M.; Hoeppner, A.; Mulnaes, D.; Buckel, W.; Gohlke, H.; Smits, S.H.; Bremer, E.; J. Mol. Biol. 426, 586-600 (2014)

Data extracted from this reference:

Crystallization (Commentary)
Crystallization (Commentary)
Organism
structural comparison, molecular dynamics simulations, and site-directed mutagenesis suggest the positioning of the iron, ectoine, and 2-oxoglutarate ligands in close proximity to each other and with a spatial orientation that will allow the region-selective and stereo-specific hydroxylation of (4S)-ectoine to (4S,5S)-5-hydroxyectoine
Virgibacillus salexigens
Engineering
Protein Variants
Commentary
Organism
A163C
residue is not involved in ligand binding
Virgibacillus salexigens
A163C/S244C
residues are not involved in ligand binding
Virgibacillus salexigens
D148A
loss of activity. Residue is involved in binding of Fe2+
Virgibacillus salexigens
D148E
loss of activity. Residue is involved in binding of Fe2+
Virgibacillus salexigens
F143A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
F143W
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
F143Y
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
F242A
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
F242W
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
F242Y
3fold increase in Km value
Virgibacillus salexigens
F263A
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
F263W
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
F263Y
30% increase in Km value
Virgibacillus salexigens
F95A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
H146A
loss of activity. Residue is involved in binding of Fe2+
Virgibacillus salexigens
H248A
loss of activity. Residue is involved in binding of Fe2+
Virgibacillus salexigens
N133A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
N261A
residue is not involved in ligand binding
Virgibacillus salexigens
P198A
activity similar to wild-type, residue of loop region
Virgibacillus salexigens
Q129A
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
R131A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
R259A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
R259H
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
R259K
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
R259Q
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
S165A
3fold increase in Km value
Virgibacillus salexigens
S167A
residue is not involved in ligand binding
Virgibacillus salexigens
S205A
activity similar to wild-type, residue of loop region
Virgibacillus salexigens
S244C
residue is not involved in ligand binding
Virgibacillus salexigens
S250A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
V265A
residue is not involved in ligand binding
Virgibacillus salexigens
V265L
residue is not involved in ligand binding
Virgibacillus salexigens
V265T
residue is not involved in ligand binding
Virgibacillus salexigens
W152A
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
W152F
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
W152Y
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
5.9
-
ectoine
wild-type, pH 7.5, 32°C
Virgibacillus salexigens
6.8
-
ectoine
mutant S205A, pH 7.5, 32°C
Virgibacillus salexigens
7.1
-
ectoine
mutant P198A, pH 7.5, 32°C
Virgibacillus salexigens
8
-
ectoine
mutant F263Y, pH 7.5, 32°C
Virgibacillus salexigens
17.3
-
ectoine
mutant S165A, pH 7.5, 32°C
Virgibacillus salexigens
19.6
-
ectoine
mutant F242Y, pH 7.5, 32°C
Virgibacillus salexigens
Organism
Organism
UniProt
Commentary
Textmining
Virgibacillus salexigens
Q2TDY4
-
-
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
Substrate Product ID
ectoine + 2-oxoglutarate + O2
-
740868
Virgibacillus salexigens
5-hydroxyectoine + succinate + CO2
-
-
-
?
Crystallization (Commentary) (protein specific)
Crystallization
Organism
structural comparison, molecular dynamics simulations, and site-directed mutagenesis suggest the positioning of the iron, ectoine, and 2-oxoglutarate ligands in close proximity to each other and with a spatial orientation that will allow the region-selective and stereo-specific hydroxylation of (4S)-ectoine to (4S,5S)-5-hydroxyectoine
Virgibacillus salexigens
Engineering (protein specific)
Protein Variants
Commentary
Organism
A163C
residue is not involved in ligand binding
Virgibacillus salexigens
A163C/S244C
residues are not involved in ligand binding
Virgibacillus salexigens
D148A
loss of activity. Residue is involved in binding of Fe2+
Virgibacillus salexigens
D148E
loss of activity. Residue is involved in binding of Fe2+
Virgibacillus salexigens
F143A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
F143W
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
F143Y
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
F242A
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
F242W
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
F242Y
3fold increase in Km value
Virgibacillus salexigens
F263A
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
F263W
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
F263Y
30% increase in Km value
Virgibacillus salexigens
F95A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
H146A
loss of activity. Residue is involved in binding of Fe2+
Virgibacillus salexigens
H248A
loss of activity. Residue is involved in binding of Fe2+
Virgibacillus salexigens
N133A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
N261A
residue is not involved in ligand binding
Virgibacillus salexigens
P198A
activity similar to wild-type, residue of loop region
Virgibacillus salexigens
Q129A
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
R131A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
R259A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
R259H
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
R259K
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
R259Q
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
S165A
3fold increase in Km value
Virgibacillus salexigens
S167A
residue is not involved in ligand binding
Virgibacillus salexigens
S205A
activity similar to wild-type, residue of loop region
Virgibacillus salexigens
S244C
residue is not involved in ligand binding
Virgibacillus salexigens
S250A
loss of activity. Residue is involved in binding of 2-oxoglutarate
Virgibacillus salexigens
V265A
residue is not involved in ligand binding
Virgibacillus salexigens
V265L
residue is not involved in ligand binding
Virgibacillus salexigens
V265T
residue is not involved in ligand binding
Virgibacillus salexigens
W152A
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
W152F
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
W152Y
loss of activity. Residue is involved in binding of ectoine
Virgibacillus salexigens
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
5.9
-
ectoine
wild-type, pH 7.5, 32°C
Virgibacillus salexigens
6.8
-
ectoine
mutant S205A, pH 7.5, 32°C
Virgibacillus salexigens
7.1
-
ectoine
mutant P198A, pH 7.5, 32°C
Virgibacillus salexigens
8
-
ectoine
mutant F263Y, pH 7.5, 32°C
Virgibacillus salexigens
17.3
-
ectoine
mutant S165A, pH 7.5, 32°C
Virgibacillus salexigens
19.6
-
ectoine
mutant F242Y, pH 7.5, 32°C
Virgibacillus salexigens
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ID
ectoine + 2-oxoglutarate + O2
-
740868
Virgibacillus salexigens
5-hydroxyectoine + succinate + CO2
-
-
-
?
Other publictions for EC 1.14.11.55
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)
741719
Tao
Ectoine and 5-hydroxyectoine ...
Virgibacillus halodenitrificans, Virgibacillus halodenitrificans PDB-F2
Appl. Microbiol. Biotechnol.
100
6779-6789
2016
-
-
-
-
-
-
-
-
-
-
-
2
-
2
-
-
-
-
-
-
-
-
2
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-
1
-
-
1
-
-
742383
Widderich
Strangers in the archaeal wor ...
Nitrosopumilus maritimus
Environ. Microbiol.
18
1227-1248
2016
-
-
1
-
-
-
-
-
-
-
1
1
-
7
-
-
1
-
-
-
-
-
1
1
1
-
-
-
-
-
-
-
-
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-
1
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-
1
1
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-
1
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-
-
-
1
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
741466
Yang
Crystallization and X-ray diff ...
Bacillus pseudofirmus, Bacillus pseudofirmus OF4
Wei Sheng Wu Xue Bao
55
1468-1474
2015
-
-
1
1
-
-
-
-
-
-
-
-
-
5
-
-
-
-
-
-
-
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-
1
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1
-
1
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-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
739813
Hoeppner
Overexpression, crystallizatio ...
Sphingopyxis alaskensis
Acta Crystallogr. Sect. F
70
493-496
2014
-
-
1
1
-
-
-
-
-
1
-
-
-
2
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
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-
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-
1
-
1
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1
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-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
740729
Hoeppner
Crystal structure of the ectoi ...
Sphingopyxis alaskensis, Sphingopyxis alaskensis DSM 13593
J. Biol. Chem.
289
29570-29583
2014
-
-
-
1
7
-
-
2
-
-
2
-
-
5
-
-
-
-
-
-
-
-
2
1
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
7
-
-
-
-
2
-
-
2
-
-
-
-
-
-
-
-
-
2
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
740868
Widderich
Molecular dynamics simulations ...
Virgibacillus salexigens
J. Mol. Biol.
426
586-600
2014
-
-
-
1
36
-
-
6
-
-
-
-
-
3
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
36
-
-
-
-
6
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
741304
Widderich
Biochemical properties of ecto ...
Acidiphilium cryptum, Alkalilimnicola ehrlichii, Halomonas elongata, Halomonas elongata DSM 2581, Paenibacillus lautus, Pseudomonas stutzeri, Pseudomonas stutzeri A1501, Sphingopyxis alaskensis, Sphingopyxis alaskensis DSM 13593, Virgibacillus salexigens
PLoS ONE
9
e93809
2014
-
-
-
1
-
-
7
14
-
21
7
20
-
17
-
-
-
-
-
-
-
-
20
7
8
7
7
-
7
7
7
-
-
-
7
-
-
-
-
-
1
-
-
-
7
-
14
-
21
7
20
-
-
-
-
-
-
-
-
20
7
7
7
-
7
7
7
-
7
-
-
-
-
7
7
743087
Widderich
Molecular dynamics simulation ...
Virgibacillus salexigens
J. Mol. Biol.
426
586-600
2014
-
-
1
-
36
-
-
14
-
1
-
1
-
3
-
-
-
-
-
-
-
-
1
-
1
-
-
-
-
-
-
-
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-
-
-
-
1
-
-
36
-
-
-
-
14
-
1
-
1
-
-
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-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
740784
Eilert
Synthesis and release of the b ...
Halomonas elongata, Halomonas elongata DSM 2581
J. Biotechnol.
167
85-93
2013
-
1
-
-
-
-
-
-
-
-
-
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-
3
-
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-
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-
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1
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1
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-
741265
Reuter
Synthesis of 5-hydroxyectoine ...
Virgibacillus salexigens
PLoS ONE
5
e10647
2010
-
-
1
1
-
-
-
-
-
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-
-
5
-
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-
-
-
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1
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1
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-
739899
Bursy
Synthesis and uptake of the co ...
Streptomyces coelicolor, Streptomyces coelicolor A3(2)
Appl. Environ. Microbiol.
74
7286-7296
2008
-
-
-
-
-
-
1
2
-
1
1
-
-
43
-
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1
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1
-
4
1
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1
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1
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1
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2
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1
1
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1
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1
-
4
1
1
-
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-
1
-
-
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-
1
1
-
-
-
740684
Bursy
Osmotically induced synthesis ...
Virgibacillus salexigens
J. Biol. Chem.
282
31147-31155
2007
-
-
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1
2
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1
3
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6
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1
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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
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1
3
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1
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1
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2
1
1
-
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1
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-
-
1
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-
1
-
-
739896
Prabhu
Functional expression of the e ...
Streptomyces anulatus
Appl. Environ. Microbiol.
70
3130-3132
2004
-
1
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3
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1
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1
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1
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1
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