Information on EC 1.14.18.6 - 4-hydroxysphinganine ceramide fatty acyl 2-hydroxylase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.14.18.6
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RECOMMENDED NAME
GeneOntology No.
4-hydroxysphinganine ceramide fatty acyl 2-hydroxylase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
a phytoceramide + 2 ferrocytochrome b5 + O2 + 2 H+ = a (2'R)-2'-hydroxyphytoceramide + 2 ferricytochrome b5 + H2O
show the reaction diagram
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
fatty acid alpha-oxidation III
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sphingolipid biosynthesis (yeast)
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SYSTEMATIC NAME
IUBMB Comments
(4R)-4-hydroxysphinganine ceramide,ferrocytochrome-b5:oxygen oxidoreductase (fatty acyl 2-hydroxylating)
The enzyme, characterized from yeast and mammals, catalyses the hydroxylation of carbon 2 of long- or very-long-chain fatty acids attached to (4R)-4-hydroxysphinganine during de novo ceramide synthesis. The enzymes from yeast and from mammals contain an N-terminal cytochrome b5 domain that acts as the direct electron donor to the desaturase active site. The newly introduced 2-hydroxyl group has R-configuration. cf. EC 1.14.18.7, dihydroceramide fatty acyl 2-hydroxylase.
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
QO3529
FA2H/Scs7p belongs to a superfamily of integral membrane di-iron-containing enzymes that hydroxylate or desaturate lipid-based substrates via a reaction mechanism that is dependent on NADH and oxygen
physiological function
additional information
QO3529
important role of the dimetal-binding histidines in catalysis, residues Tyr322 and Asp323 are critical determinants involved in the hydroxylase reaction. Molecular dynamics simulations, structure-function analysis, and generation of a model of ceramide binding to enzyme Scs7p
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
a phytoceramide + 2 ferrocytochrome b5 + O2 + 2 H+
a (2'R)-2'-hydroxyphytoceramide + 2 ferricytochrome b5 + H2O
show the reaction diagram
monohexosylceramide + 2 ferrocytochrome b5 + O2 + 2 H+
2'-hydroxymonohexosylceramide + 2 ferricytochrome b5 + H2O
show the reaction diagram
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?
palmitic acid + 2 ferrocytochrome b5 + O2 + 2 H+
(R)-2-hydroxypalmitic acid + 2 ferricytochrome b5 + H2O
show the reaction diagram
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FA2H is stereospecific for the production of (R)-enantiomers
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?
phytoceramide + ferrocytochrome b5 + O2 + H+
(2'R)-2'-hydroxyphytoceramide + ferricytochrome b5 + H2O
show the reaction diagram
QO3529
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?
additional information
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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
a phytoceramide + 2 ferrocytochrome b5 + O2 + 2 H+
a (2'R)-2'-hydroxyphytoceramide + 2 ferricytochrome b5 + H2O
show the reaction diagram
QO3529
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cytochrome b5
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Zn2+
QO3529
the enzyme contains two zinc atoms; the enzyme contains two zinc atoms coordinated by the side chains of 10 highly conserved histidines within a dimetal center located near the plane of the cytosolic membrane. The dimetal-binding site is located within the catalytic cap domain and is occupied by zinc atoms coordinated by the highly conserved histidine side chains from four histidine box motifs. Molecular dynamics simulations, allows modeling of a ceramide substrate below the dimetal center of scScs7p
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Bax inhibitor-1
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from Arabidopsis thaliana, BI-1 (AtBI-1) protein interacts with the enzyme. Bax inhibitor-1 is a widely conserved cell death suppressor localized in the endoplasmic reticulum membrane. AtBI-1 binds cytochrome b5-like domain-containing proteins like fatty acid 2-hydroxylase 1
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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FA2H is expressed in cultured human keratinocytes and human epidermis, with FA2H expression and fatty acid 2-hydroxylase activity increasing with differentiation
Manually annotated by BRENDA team
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FA2H is expressed in cultured human keratinocytes and human epidermis, with FA2H expression and fatty acid 2-hydroxylase activity increasing with differentiation
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
QO3529
an integral membrane protein, the Scs7p core is composed of a helical catalytic cap domain that sits atop four transmembrane helices that anchor the enzyme in the endoplasmic reticulum
Manually annotated by BRENDA team
additional information
QO3529
modeling of secondary structural elements of the scScs7p hydroxylase domain placed within a model lipid bilayer, overview
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Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
70000
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x * 71800, calculated, x * 70000, SDS-PAGE
71800
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x * 71800, calculated, x * 70000, SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 71800, calculated, x * 70000, SDS-PAGE
additional information
QO3529
the core structure is composed of a helical catalytic cap domain that sits atop four transmembrane helices that anchor the enzyme in the endoplasmic reticulum. The dimetal-binding site is located within the catalytic cap domain and is occupied by zinc atoms coordinated by the highly conserved histidine side chains from four histidine box motifs
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
microbatch-under-oil method, using 20% polyethylene glycol 3350 (w/v), 100 mM HEPES, pH 7.0, 300 mM ammonium sulfate; purified recombinant wild-type scScs7p and mutant DELTA95scScs7p, microbatch-under-oil method, micxing of 0.002 ml of protein solution at 2.5 or 3.3 mg/ml with 0.002 ml of crystallization solution containing 10% PEG 2000, 100 mM sodium citrate, pH 5.6, and 50 mM ammonium sulfate, or 20% w/v PEG 3350, 100 mM HEPES, pH 7.0, and 300 mM ammonium sulfate, and then covering the drop with 0.05 ml of mineral oil, 2-7 days, 14°C, soaking in heavy atom solution saturated with K2HgI4, X-ray diffraction structure determination and analysis at 2.6-6.0 A resolution
QO3529
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
immunoglobulin G-Sepharose 6 resin column chromatography and Superdex 200 gel filtration; recombinant tagged wild-type scScs7p and DELTA95scScs7p from Saccharomyces cerevisiae strain BJ5460 membranes by solubilization with beta-octyl glucoside, ultracentrifugation, affinity/immunoaffinity chromatography, and tag cleavage through rhinovirus 3C protease, which is removed by adding His-Select cobalt resin, followed by ultrafiltration and gel filtration
QO3529
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Saccharomyces cerevisiae strain BJ5460; gene SCS7, recombinant overexpression of ZZ/His6/MBP-tagged wild-type scScs7p and DELTA95scScs7p from plasmid pSGP46 in Saccharomyces cerevisiae strain BJ5460, recombinant expression of point mutants in enzyme-deficient Saccharomyces cerevisiae strain
QO3529
expression in CHO cell
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expression in CHO cells
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expression in HCT116 cells
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
DELTA9-tetrahydrocannabinol together with induced levels of peroxisome proliferator-activated receptor alpha significantly stimulate the expression of fatty acid 2 hydroxylase
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differentiation-dependent up-regulation of ceramide synthesis and fatty acid elongation is accompanied by up-regulation of FA2H
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fatty acid 2-hydroxylase (FA2H) is upregulated in vitro by nimodipine, a component of myelin synthesis. Nimodipine but not nifedipine increases FA2H protein levels and also significantly increases mRNA levels of FA2H in both undifferentiated and differentiated Neuro2a cells. Osmotic (NaCl) stress increases FA2H mRNA expression, synergistically with nifedipine-induced stress, overview
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nimodipine promotes expression of fatty acid 2-hydroxylase in a surgical stress model based on Neuro2a cells
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treatment of D6P2T Schwannoma cells with dibutyryl-cAMP induces exit from the cell cycle with concomitant upregulation of FA2H
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D323A
QO3529
site-directed mutagenesis, the mutation results in partial loss of syringomycin E sensitivity
H173A
QO3529
site-directed mutagenesis, the mutation results in partial loss of syringomycin E sensitivity
H244A
QO3529
site-directed mutagenesis, the mutation results in major loss of syringomycin E sensitivity
H249A
QO3529
site-directed mutagenesis, the mutation results in partial loss of syringomycin E sensitivity
H264A
QO3529
site-directed mutagenesis, the mutation results in unaltered syringomycin E sensitivity compared to the wild-type
H268A
QO3529
site-directed mutagenesis, the mutation results in partial loss of syringomycin E sensitivity
H271A
QO3529
site-directed mutagenesis, the mutation results in major loss of syringomycin E sensitivity
H272A
QO3529
site-directed mutagenesis, the mutation results in major loss of syringomycin E sensitivity
H326A
QO3529
site-directed mutagenesis, the mutation results in major loss of syringomycin E sensitivity
H330A
QO3529
site-directed mutagenesis, the mutation results in complete loss of syringomycin E sensitivity
H331A
QO3529
site-directed mutagenesis, the mutation results in unaltered syringomycin E sensitivity compared to the wild-type
H345A
QO3529
site-directed mutagenesis, the mutation results in major loss of syringomycin E sensitivity
H348A
QO3529
site-directed mutagenesis, the mutation results in major loss of syringomycin E sensitivity
H349A
QO3529
site-directed mutagenesis, the mutation results in major loss of syringomycin E sensitivity
Y319A
QO3529
site-directed mutagenesis, the mutation results in unaltered syringomycin E sensitivity compared to the wild-type
Y322A
QO3529
site-directed mutagenesis, the mutation results in major loss of syringomycin E sensitivity
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine