Information on EC 4.4.1.16 - selenocysteine lyase

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea

EC NUMBER
COMMENTARY hide
4.4.1.16
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RECOMMENDED NAME
GeneOntology No.
selenocysteine lyase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
L-selenocysteine + reduced acceptor = selenide + L-alanine + acceptor
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
alpha,beta-elimination
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-
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cleavage of C-Se bond
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elimination of chloride
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-
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elimination of H2Se
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-
-
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elimination of SO2
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-
-
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oxidative elimination of NH3
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-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
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Selenocompound metabolism
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selenocysteine biosynthesis
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SYSTEMATIC NAME
IUBMB Comments
L-selenocysteine selenide-lyase (L-alanine-forming)
A pyridoxal-phosphate protein. Dithiothreitol or 2-mercaptoethanol can act as the reducing agent in the reaction. The enzyme from animals does not act on cysteine, serine or chloroalanine [1,3], while the enzyme from bacteria shows activity with cysteine (cf. EC 2.8.1.7, cysteine desulfurase) [2].
CAS REGISTRY NUMBER
COMMENTARY hide
82047-76-5
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
L. Czern., wild type and transgenic line overexpressing selenocysteine lyase of Mus musculus
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
strain IRC 0070
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Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
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SwissProt
Manually annotated by BRENDA team
C57BL/6
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Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
Sprague-Dawley
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Manually annotated by BRENDA team
Wistar
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
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disruption of the selenocysteine lyase-mediated selenium recycling pathway leads to metabolic syndrome in mice affecting hepatic glucose and lipid homeostasis. Mice lacking the enzyme and raised on an Se-adequate diet exhibit hyperinsulinemia, hyperleptinemia, glucose intolerance, and hepatic steatosis, with increased hepatic oxidative stress, but maintain selenoprotein levels and circulating Se status. Insulin challenge of enyme KO mice results in attenuated Akt phosphorylation but does not decrease phosphorylation levels of AMP kinase alpha. Upon dietary Se restriction, enzyme KO animals develop several characteristics of metabolic syndrome, such as obesity, fatty liver, and hypercholesterolemia, with aggravated hyperleptinemia, hyperinsulinemia, and glucose intolerance. Hepatic glutathione peroxidase 1 and selenoprotein S production and circulating selenoprotein P levels are significantly diminished. Enzyme disruption increases the levels of insulin-signaling inhibitor insulin signaling inhibitor protein phosphatase 1B
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
beta-chloro-L-alanine
NH3 + pyruvate + chloride
show the reaction diagram
D-cysteine + H2O
sulfide + NH3 + pyruvate
show the reaction diagram
L-cysteine sulfinic acid + reduced acceptor
SO2 + L-alanine + acceptor
show the reaction diagram
L-selenocysteine + reduced acceptor
L-alanine + selenium + acceptor
show the reaction diagram
L-selenocysteine + reduced acceptor
selenide + L-alanine + acceptor
show the reaction diagram
selenomethionine reduced acceptor
methylselenole + selenide
show the reaction diagram
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
L-selenocysteine + reduced acceptor
selenide + L-alanine + acceptor
show the reaction diagram
additional information
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
pyridoxal 5'-phosphate
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
KCl
0-0.5 M KCl gives optimal activities at around 55-60°C.When the KCl concentration is increased to 2.5-3 M, this optimum temperature shifts to between 70-75°C
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
beta-chloro-L-alanine
DTT
activity is strongly dependent on the presence of DTT, with activity increasing up to 46% when the reductant is present in the reaction mixture. Concentrations higher than 5 mM cause an inhibitory effect
hydroxylamine
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almost completely reversible by dialysis against pyridoxal 5'-phosphate
iodoacetamide
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1 mM, complete inactivation within 3 h
iodoacetic acid
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1 mM, complete inactivation within 3 h
L-cysteine
L-cysteine methyl ester
major urinary protein 1
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about 60% remaining relative enzyme activity
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major urinary protein 2
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about 60% remaining relative enzyme activity
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N-ethylmaleimide
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1 mM, complete inactivation within 3 h
NaBH4
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
alpha-Keto acids
dithiothreitol
activity is strongly dependent on the presence of dithiothreitol, with activity increasing up to 46% when the reductant is present in the reaction mixture. Concentrations higher than 5 mM cause an inhibitory effect
pyruvate
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3.1
beta-chloro-L-alanine
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pH 7.2
8.6 - 30
L-cysteine sulfinate
0.2 - 9.9
L-selenocysteine
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.35
L-cysteine sulfinate
Mus musculus
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pH 9.0, 37°C
46
L-selenocysteine
Mus musculus
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pH 9.0, 37°C
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.5
beta-chloro-L-alanine
0.65 - 9.6
L-cysteine
0.6
L-cysteine methyl ester
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.6
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purified enzyme, pH 7.5, 37°C
6.47
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purified enzyme
29
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purified enzyme, pH 7.4
37
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purified enzyme, pH 8.5, 37°C
200
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knockdown of Nfs-like proteins, activity measured in mitochondrial fractions
300
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knockdown of selenocysteine lyase, activity measured in mitochondrial fractions
600
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activity measured in mitochondrial fractions
1000
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knockdown of selenocysteine lyase, activity measured in cytosolic fractions
1200
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knockdown of Nfs-like proteins, activity measured in cytosolic fractions
1500
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activity measured in cytosolic fractions
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 7.5
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7.5
maximal activity is obtained in 50 mM phosphate buffer, pH 7.5, 2 M KCl at 65°C
8.5
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assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 10
pH 6.5: about 45% of maximal activity, pH 10.0: about 45% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
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assay at
55 - 60
0-0.5 M KCl gives optimal activities at around 55-60°C.When the KCl concentration is increased to 2.5-3 M, this optimum temperature shifts to 70-75°C
70 - 75
0-0.5 M KCl gives optimal activities at around 55-60°C.When the KCl concentration is increased to 2.5-3 M, this optimum temperature shifts to 70-75°C
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
43000
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SDS-PAGE
44440
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calculated from amino acid sequence
46800
2 * 46800, calculated from sequence
47000
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SDS-PAGE
47200
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calculated from amino acid sequence
48000
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SDS-PAGE
60200
2 * 60200, SDS-PAGE
63000
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HPLC gel filtration
85000
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sedimentation equilibrium centrifugation
88000
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gel filtration
93000
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gel filtration
105000
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gel filtration
110000
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method
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purified enzyme in dimeric and tetrameric forms, sitting drop vapour diffusion method, 100 nl of 17 mg/ml protein solution with 100 nl of well solution 1 containing 100 mM HEPES, pH 6.7, and 10% PEG 6000, or well solution 2 containing 50 mM HEPES, pH 8.1, 200 mM ammonium nitrate and 25% PEG 3350, 20°C, 3 days or 7 days, respectively, X-ray diffraction structure determination and analysis of the two crystals forms at 2.1 A and 1.8 A resolution, respectively, modeling of dimeric enzyme complexed with pyridoxal 5'-phosphate
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
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50% of activity lost within 30 min
80
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purified enzyme, pH 8.5, 2 min, inactivation
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
1% bovine serum albumin, 20% sucrose or 20% glycerol protects against inactivation, highly diluted samples sensitive to freezing
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labile during purification, 0.1% 2-mercaptoethanol, 1 mM dithiothreitol or 1 M sucrose stabilizes activity
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, 1 month, 60% loss of activity
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-20°C, 20% sucrose, stable
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-80°C, purified enzyme, stable for several weeks
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
overexpressed protein
recombiant N-terminally His6-tagged wild-type and mutant enyzmes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration, followed by tag cleavage through TEV protease, another step of nickel affinity chromatography, and dialysis
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recombinant N-terminally His6-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration, followed by tag cleavage through TEV protease, another step of nickel affinity chromatography, and dialysis
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Brassica juncea using Agrobacterium tumefaciens to introduce genetic sequence, transformation results in enhanced enzyme activity but decreased Se tolerance
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expressed in chloroplasts and cytosol of Arabidopsis sp., enhanced Se tolerance when enzyme is expressed in cytosol, decreased Se tolerance when expressed in chloroplasts
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expressed in Escherichia coli BL21(DE3)
expressed in Escherichia coli JM109
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expression in Escherichia coli
expression in Escherichia coli into the pGEM-T Easy vector for sequencing, 5’ constructs with deletions in the promoter region are subcloned into the promoterless luciferase expression vector
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expression in Escherichia coli rosetta
recombinant expression in Escherichia coli strain BL21(DE3)
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recombinant expression of GFP-tagged enzyme in spermatids of mature testes of transgenic mice
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recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
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sequence comparisons, recombiant expression of N-terminally His6-tagged wild-type and mutant enyzmes in Escherichia coli strain BL21(DE3)
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
presence or absence of selenoproteins does not affect enzyme expression in HeLa cells
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C364A
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increased activity towards L-selenocysteine and L-cysteine sulfinate, activity toward L-cysteine was completely abolished
H123A
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only 1% of activity remaining compared to wild type, contributes to stabilization of pyridoxal 5'-phospahte
H55A
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no effect on activity
R379A
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less than 1% of activity remaining compared to wild type
D146K
site-directed mutagenesis, the mutant variant shows loss of selenocysteine specificity and appearance of cysteine desulfurase activity
D146K/H389T
site-directed mutagenesis,the mutant variant shows loss of selenocysteine specificity and appearance of cysteine desulfurase activity to a higher extent
H389T
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site-directed mutagenesis, the mutant variant shows loss of selenocysteine specificity and appearance of cysteine desulfurase activity
V256S
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site-directed mutagenesis, the mutant variant shows loss of selenocysteine specificity and appearance of cysteine desulfurase activity
C375A
mutation on the catalytically essential Cys-375 residue
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
agriculture
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Selenium contamination is one of the most serious problems in agriculture in Se-rich regions, cpSL transgenic plants can be used for Se phytoremediation under field conditions. Selenium accumulation is significantly enhanced in the shoots of the cpSL transgenic plants compared to wild typ. cpSL plants are more tolerant to the toxic sediment-soil than the wild type
degradation
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transgenic plants could be used for decontamination of high Se soil or water
medicine
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