Information on EC 2.3.1.65 - bile acid-CoA:amino acid N-acyltransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.3.1.65
-
RECOMMENDED NAME
GeneOntology No.
bile acid-CoA:amino acid N-acyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
choloyl-CoA + glycine = CoA + glycocholate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acyl group transfer
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-
-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
bile acid biosynthesis, neutral pathway
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Metabolic pathways
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Primary bile acid biosynthesis
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Taurine and hypotaurine metabolism
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bile acid biosynthesis, neutral pathway
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SYSTEMATIC NAME
IUBMB Comments
choloyl-CoA:glycine N-choloyltransferase
Also acts on CoA derivatives of other bile acids. Taurine and 2-fluoro-beta-alanine can act as substrates, but more slowly [4]. The enzyme can also conjugate fatty acids to glycine and can act as a very-long-chain acyl-CoA thioesterase [7]. Bile-acid---amino-acid conjugates serve as detergents in the gastrointestinal tract, solubilizing long chain fatty acids, mono- and diglycerides, fat-soluble vitamins and cholesterol [4]. This is the second enzyme in a two-step process leading to the conjugation of bile acids with amino acids; the first step is the conversion of bile acids into their acyl-CoA thioesters, which is catalysed by EC 6.2.1.7, cholate---CoA ligase.
CAS REGISTRY NUMBER
COMMENTARY hide
74506-32-4
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
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BAT restores bile acid contents in septic livers, overview
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
chenodeoxycholoyl-CoA + glycine
CoA + glycochenodeoxycholate
show the reaction diagram
chenodeoxycholoyl-CoA + taurine
CoA + taurochenodeoxycholate
show the reaction diagram
choloyl-CoA + 2-fluoro-beta-alanine
?
show the reaction diagram
-
-
-
?
choloyl-CoA + 2-fluoro-beta-alanine
CoA + 2-fluoro-beta-alanylcholate
show the reaction diagram
-
2-fluoro-beta-alanine is an excellent substrate like taurine
-
-
?
choloyl-CoA + aminomethanesulfonic acid
?
show the reaction diagram
-
-
-
?
choloyl-CoA + beta-alanine
CoA + beta-alanylcholate
show the reaction diagram
choloyl-CoA + D-alpha-alanine
CoA + D-alpha-alanylcholate
show the reaction diagram
-
about 10% of the rate with glycine
-
-
?
choloyl-CoA + glycine
CoA + glycocholate
show the reaction diagram
choloyl-CoA + taurine
CoA + taurocholate
show the reaction diagram
deoxycholoyl-CoA + aminomethanesulfonate
CoA + aminomethanesulfonyldeoxycholate
show the reaction diagram
-
-
-
-
?
deoxycholoyl-CoA + glycine
CoA + glycodeoxycholate
show the reaction diagram
deoxycholoyl-CoA + taurine
CoA + taurodeoxycholate
show the reaction diagram
lithocholoyl-CoA + glycine
CoA + glycolithocholate
show the reaction diagram
lithocholoyl-CoA + taurine
CoA + taurolithocholate
show the reaction diagram
trihydroxycholestanoyl-CoA + glycine
CoA + ?
show the reaction diagram
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very low activity
-
-
?
ursodeoxycholoyl-CoA + glycine
CoA + glycoursodeoxycholate
show the reaction diagram
-
24.8% of activity with choloyl-CoA
-
-
?
ursodeoxycholoyl-CoA + taurine
CoA + tauroursodeoxycholate
show the reaction diagram
-
35.3% of activity with choloyl-CoA
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-
?
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
choloyl-CoA + glycine
CoA + glycocholate
show the reaction diagram
choloyl-CoA + taurine
CoA + taurocholate
show the reaction diagram
additional information
?
-
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4-hydroxynonenal
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inactivated by 4HNE in a dose-dependent manner. The active-site His (His362) dose-dependently forms a 4-hydroxynonenal adduct, contributing to loss of activity
5,5'-dithiobis(2-nitrobenzoic acid)
chenodeoxycholate
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-
cholate
cholesterol
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Conjugated bile acids
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efficient competitive inhibitors of cholyl-CoA binding
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deoxycholate
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-
glycocholate
glycodeoxycholate
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N-ethylmaleimide
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10 min, at 0.2 mM: 90% loss of activity, preincubation with cholyl-CoA before NEM-treatment protects
NaCl
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above 50 mM
p-mercuribenzoate
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reversible by dithioerythritol, substrates protect, serine and alanine protect
taurocholate
taurodeoxycholate
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-
ursodeoxycholate
-
-
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
EDTA
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2fold activation, lowers the Km-value
L-cysteine
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2fold activation
Pectin
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dietary pectin relative to cellulose activates, especially the glycine-dependent activity
reduced glutathione
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2fold activation
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2
2-fluoro-beta-alanine
37°C, pH 8.4
0.8
aminomethanesulfonate
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+ 0.05 mM deoxycholoyl-CoA
2
aminomethanesulfonic acid
37°C, pH 8.4
175
beta-Alanine
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-
0.0072 - 0.087
choloyl-CoA
3.2 - 40
glycine
0.6 - 6.7
taurine
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
additional information
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.038
glycocholate
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0635
activity measured with taurine as substrate
0.42
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taurine
0.43
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glycine
1.041
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activity with choloyl-CoA anf glycine as substrates
1.99
-
taurine
3.88
-
glycine
35.2
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taurine
43 - 59
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glycine
46.9
-
glycine
additional information
-
-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5
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taurine-dependent activity
7.2
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glycine-dependent activity
additional information
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.6 - 8.2
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taurine: about half-maximal activity at pH 5.2 and pH 8.2, glycine: 80% of maximal activity at pH 8.2
6 - 9
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about 70% of maximal activity at pH 6.0, about half-maximal activity at pH 9.0, inactivation rate is similar for both taurine and glycine
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45000
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x * 46525, calculated from the amino acid sequence, x * 45000, recombinant enzyme, SDS-PAGE
45700
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gel filtration
46525
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x * 46525, calculated from the amino acid sequence, x * 45000, recombinant enzyme, SDS-PAGE
47000
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gel filtration
50000
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x * 50000, SDS-PAGE
50900
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1 * 50900, SDS-PAGE
52000
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2 * 52000, SDS-PAGE
63000 - 65000
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100000
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gel filtration
118000
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sucrose density gradient centrifugation
additional information
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amino acid composition
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
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2 * 52000, SDS-PAGE
monomer
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1 * 50900, SDS-PAGE
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6
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phosphate buffer, t1/2: 2 h
487368
9
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Tris-buffer, t1/2: 3 h
487368
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45
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stable below
57
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10 min, about 70% loss of activity
60
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inactivation within 10 min
additional information
-
taurocholate stabilizes against temperature denaturation
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
freezing inactivates
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glycerol, 40% v/v, stabilizes during storage
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high ionic strength stabilizes
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unstable enzyme
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, in the presence of 40% v/v glycerol
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-70°C, several weeks, stable
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0° to 20°C, several h, stable
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0°C, 50 mM phosphate buffer, pH 7.8, with 0.5 M NaCl, 2 mM dithioerythritol, 1 month, 25% loss of activity
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
900fold
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about 100fold
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partial purification of recombinant enzyme, expressed in Escherichia coli DH5alpha
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partial, 200fold
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cDNA encoding enzyme is cloned, sequenced and expressed in Escherichia coli DH5alpha, the open-reading frame codes for a 420 amino acid protein, Baat gene is mapped to mouse chromosome 4, single gene encodes enzyme
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expression in Escherichia coli
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expression in Sf9 cells
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expression in Sf9 cells. Initial attempts to express rBAT in Escherichia coli with vectors such as pMAL-p2 and pKK233-2 are unsuccessful due to proteolytic degradation of the expressed protein
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expression of GFP-tagged enzyme in HeLa cells, which is exclusively located in the cytosol, high mobility of the GFP-fusion proteins in the cytosol and absence of peroxisomal escape
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expression of wild-type and mutant enzymes
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gene/full-length cDNA is cloned and expressed in Escherichia coli XL1-Blue or BL21
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when expressed in fibroblasts, GFP-tagged hBAAT localizes to the cytosol
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
dexamethasone-induced BAT expression reversing the nuclear retinoid X receptor alpha reduction in septic livers, overview
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C235A
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activity is less than 0.4% of the wild-type activity
C372A
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mutant with low enzyme activity
D326A
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activity is less than 0.4% of the wild-type activity
D328A
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inactive enzyme
H362A
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inactive enzyme
H362Q
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activity is less than 0.4% of the wild-type activity
additional information
-
the cytosolic enzyme localization is due to a carboxyterminal non-consensus peroxisomal targeting signal, SQL, since mutation of the SQL to SKL results in BAAT being efficiently imported into peroxisomes