BRENDA - Enzyme Database show
show all sequences of 1.14.14.25

Expression of human cytochrome P450 46A1 in Escherichia coli: effects of N- and C-terminal modifications

Mast, N.; Andersson, U.; Nakayama, K.; Bjorkhem, I.; Pikuleva, I.A.; Arch. Biochem. Biophys. 428, 99-108 (2004)

Data extracted from this reference:

Cloned(Commentary)
Commentary
Organism
expression in Escherichia coli, wild-type and trucation mutatants. All four mutants lack the N-terminal transmembrane region (residues 3–27), and, in addition, DELTA46A1 has a 4 His-tag fused to the C-terminus, HDELTA46A1 has the N-terminal 4 His-tag, HDELTA46A1DELTA has a 4 His-tag at the N-terminus and does not contain a proline-rich region at the C-terminus (residues 494–499), and DELTA46A1DELTA lacks the C-terminal proline-rich region
Homo sapiens
Engineering
Amino acid exchange
Commentary
Organism
additional information
four truncation mutants: all lack the N-terminal transmembrane region (residues 3 –27), and, in addition, DELTA46A1 has a 4 His-tag fused to the C-terminus, HDELTA46A1 has the N-terminal 4 His-tag, HDELTA46A1DELTA has a 4 His-tag at the N-terminus and does not contain a proline-rich region at the C-terminus (residues 494 –499), and DELTA46A1DELTA lacks the C-terminal proline-rich region. Truncated enzymes have moderately decreased catalytic efficiencies for either cholesterol or 24S-hydroxycholesterol or both, whereas their substrate-binding constants are either unchanged or decreased 2fold.The two forms, DELTA64A1DELTA and HDELTA46A1DELTA both lacking the C-terminal proline-rich region are good candidates for future crystallographic studies because they contain only 0.3 –0.8% of high molecular weight aggregates and their catalytic efficiencies are decreased no more than 2.3fold
Homo sapiens
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
additional information
-
additional information
Km-value of truncated enzyme forms
Homo sapiens
0.0022
-
(24S)-24-hydroxycholesterol
full length enzyme
Homo sapiens
Localization
Localization
Commentary
Organism
GeneOntology No.
Textmining
cytosol
truncated enzymes are distributed at about a 1:1 ratio between the membrane fraction and the cytosol in low ionic strength buffer, when expressed in Escherichia coli
Homo sapiens
5829
-
membrane
truncated enzymes are distributed at about a 1:1 ratio between the membrane fraction and the cytosol in low ionic strength buffer, when expressed in Escherichia coli
Homo sapiens
16020
-
Molecular Weight [Da]
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
49000
-
1 * 49000, truncated enzyme form, SDS-PAGE
Homo sapiens
Organism
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
Homo sapiens
-
-
-
Purification (Commentary)
Commentary
Organism
truncated enzyme forms
Homo sapiens
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
(24S)-24-hydroxycholesterol + [reduced NADPH-hemoprotein reductase] + O2
-
657671
Homo sapiens
24,25-dihydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
-
-
?
(24S)-24-hydroxycholesterol + [reduced NADPH-hemoprotein reductase] + O2
-
657671
Homo sapiens
24,27-dihydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
-
-
?
cholesterol + [reduced NADPH-hemoprotein reductase] + H+ + O2
-
657671
Homo sapiens
(24S)-24-hydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
-
-
?
Subunits
Subunits
Commentary
Organism
monomer
1 * 49000, truncated enzyme form, SDS-PAGE
Homo sapiens
Turnover Number [1/s]
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
additional information
-
additional information
turnover-numbers of truncated enzyme forms
Homo sapiens
0.00117
-
(24S)-24-hydroxycholesterol
full length enzyme
Homo sapiens
Cofactor
Cofactor
Commentary
Organism
Structure
cytochrome P-450
-
Homo sapiens
NADPH
-
Homo sapiens
Cloned(Commentary) (protein specific)
Commentary
Organism
expression in Escherichia coli, wild-type and trucation mutatants. All four mutants lack the N-terminal transmembrane region (residues 3–27), and, in addition, DELTA46A1 has a 4 His-tag fused to the C-terminus, HDELTA46A1 has the N-terminal 4 His-tag, HDELTA46A1DELTA has a 4 His-tag at the N-terminus and does not contain a proline-rich region at the C-terminus (residues 494–499), and DELTA46A1DELTA lacks the C-terminal proline-rich region
Homo sapiens
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
cytochrome P-450
-
Homo sapiens
NADPH
-
Homo sapiens
Engineering (protein specific)
Amino acid exchange
Commentary
Organism
additional information
four truncation mutants: all lack the N-terminal transmembrane region (residues 3 –27), and, in addition, DELTA46A1 has a 4 His-tag fused to the C-terminus, HDELTA46A1 has the N-terminal 4 His-tag, HDELTA46A1DELTA has a 4 His-tag at the N-terminus and does not contain a proline-rich region at the C-terminus (residues 494 –499), and DELTA46A1DELTA lacks the C-terminal proline-rich region. Truncated enzymes have moderately decreased catalytic efficiencies for either cholesterol or 24S-hydroxycholesterol or both, whereas their substrate-binding constants are either unchanged or decreased 2fold.The two forms, DELTA64A1DELTA and HDELTA46A1DELTA both lacking the C-terminal proline-rich region are good candidates for future crystallographic studies because they contain only 0.3 –0.8% of high molecular weight aggregates and their catalytic efficiencies are decreased no more than 2.3fold
Homo sapiens
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
additional information
-
additional information
Km-value of truncated enzyme forms
Homo sapiens
0.0022
-
(24S)-24-hydroxycholesterol
full length enzyme
Homo sapiens
Localization (protein specific)
Localization
Commentary
Organism
GeneOntology No.
Textmining
cytosol
truncated enzymes are distributed at about a 1:1 ratio between the membrane fraction and the cytosol in low ionic strength buffer, when expressed in Escherichia coli
Homo sapiens
5829
-
membrane
truncated enzymes are distributed at about a 1:1 ratio between the membrane fraction and the cytosol in low ionic strength buffer, when expressed in Escherichia coli
Homo sapiens
16020
-
Molecular Weight [Da] (protein specific)
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
49000
-
1 * 49000, truncated enzyme form, SDS-PAGE
Homo sapiens
Purification (Commentary) (protein specific)
Commentary
Organism
truncated enzyme forms
Homo sapiens
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
(24S)-24-hydroxycholesterol + [reduced NADPH-hemoprotein reductase] + O2
-
657671
Homo sapiens
24,25-dihydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
-
-
?
(24S)-24-hydroxycholesterol + [reduced NADPH-hemoprotein reductase] + O2
-
657671
Homo sapiens
24,27-dihydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
-
-
?
cholesterol + [reduced NADPH-hemoprotein reductase] + H+ + O2
-
657671
Homo sapiens
(24S)-24-hydroxycholesterol + [oxidized NADPH-hemoprotein reductase] + H2O
-
-
-
?
Subunits (protein specific)
Subunits
Commentary
Organism
monomer
1 * 49000, truncated enzyme form, SDS-PAGE
Homo sapiens
Turnover Number [1/s] (protein specific)
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
additional information
-
additional information
turnover-numbers of truncated enzyme forms
Homo sapiens
0.00117
-
(24S)-24-hydroxycholesterol
full length enzyme
Homo sapiens
Other publictions for EC 1.14.14.25
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)
735877
Boussicault
CYP46A1, the rate-limiting enz ...
Homo sapiens, Mus musculus, Mus musculus C57BL/6
Brain
139
953-970
2016
-
1
2
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1
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3
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6
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9
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1
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9
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3
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4
4
-
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-
736502
Anderson
Cholesterol hydroxylase CYP46A ...
Homo sapiens
J. Biol. Chem.
291
11876-81186
2016
2
1
1
1
7
-
-
1
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
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2
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2
1
1
2
1
7
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1
1
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1
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1
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2
-
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1
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2
2
-
-
-
736072
Uto
Imidazo[1,2-a]pyridines as cho ...
Homo sapiens
Expert Opin. Ther. Pat.
25
373-377
2015
-
1
-
-
-
-
11
-
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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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11
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1
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1
1
-
-
-
735602
Fourgeux
In vivo consequences of choles ...
Rattus norvegicus, Rattus norvegicus Wistar
Biochem. Biophys. Res. Commun.
446
775-781
2014
-
-
1
-
-
-
1
-
-
-
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2
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3
-
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5
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2
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1
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1
1
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1
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2
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5
-
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2
-
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-
-
-
-
-
1
1
1
1
-
-
728617
Maioli
Is it possible to improve memo ...
Homo sapiens
PLoS ONE
8
e68534
2013
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1
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1
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5
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4
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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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4
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1
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-
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-
1
1
-
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-
712869
Liao
Quantification of cholesterol- ...
Homo sapiens
J. Proteome Res.
10
241-248
2011
-
-
1
-
-
-
-
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1
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2
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3
-
-
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6
-
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2
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1
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1
1
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1
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2
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8
-
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2
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-
-
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-
727298
Fourgeux
24S-hydroxycholesterol and cho ...
Homo sapiens
Chem. Phys. Lipids
164
496-499
2011
-
-
-
-
-
-
-
-
1
-
-
1
-
2
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2
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1
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2
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1
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1
1
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-
705008
Shafaati
The antifungal drug voriconazo ...
Homo sapiens, Mus musculus
J. Lipid Res.
51
318-323
2010
-
-
1
-
-
-
2
-
-
-
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4
-
-
1
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2
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2
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2
1
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1
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1
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2
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2
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1
1
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705825
Hudry
Adeno-associated Virus gene th ...
Homo sapiens, Mus musculus, Mus musculus APP23
Mol. Ther.
18
44-53
2010
-
-
1
-
1
-
-
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7
-
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3
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3
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2
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1
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3
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3
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2
2
-
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-
711641
Cartagena
24S-hydroxycholesterol effects ...
Homo sapiens
Brain Res.
1319
1-12
2010
-
-
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1
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1
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4
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1
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4
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1
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1
3
3
1
-
-
712120
Smiljanic
Brain injury induces cholester ...
Rattus norvegicus, Rattus norvegicus Wistar
Histochem. Cell Biol.
134
159-169
2010
-
-
-
-
-
-
-
-
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167
-
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4
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4
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1
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1
-
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712816
Tian
Increased expression of choles ...
Homo sapiens
J. Neurochem.
113
978-989
2010
-
-
1
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2
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3
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3
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1
1
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-
695874
Mast
Combined use of mass spectrome ...
Homo sapiens
Arch. Biochem. Biophys.
483
81-89
2009
-
-
1
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-
-
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1
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2
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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
1
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-
698342
Fu
Cholesterol 24-hydroxylase (CY ...
Homo sapiens
Int. J. Geriatr. Psychiatry
24
921-926
2009
-
1
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-
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2
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1
1
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-
701715
Russell
Cholesterol 24-hydroxylase: an ...
Bos taurus, Canis lupus familiaris, Cavia porcellus, Danio rerio, Equus caballus, Equus sp., Gallus gallus, Homo sapiens, Macaca mulatta, Mus musculus, Ornithorhynchus anatinus, Oryctolagus cuniculus, Pan troglodytes, Rattus norvegicus, Xenopus laevis
Annu. Rev. Biochem.
78
1017-1040
2009
-
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2
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3
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1
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17
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11
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7
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7
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2
7
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3
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1
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11
-
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7
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2
3
3
2
-
-
702028
Shafaati
Transcriptional regulation of ...
Homo sapiens, Mus musculus, Mus musculus C57B6-J
Biochem. Biophys. Res. Commun.
378
689-694
2009
-
-
-
-
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4
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5
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2
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2
-
-
703580
Koelsch
CYP46A1 variants influence Alz ...
Homo sapiens
Eur. Psychiatry
24
183-190
2009
-
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2
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1
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1
1
-
-
-
704109
Fourgeux
Primary open angle glaucoma: A ...
Homo sapiens
Invest. Ophthalmol. Vis. Sci.
50
5712-5717
2009
-
-
-
-
-
-
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2
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1
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