Information on EC 2.4.1.85 - cyanohydrin beta-glucosyltransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.4.1.85
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RECOMMENDED NAME
GeneOntology No.
cyanohydrin beta-glucosyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
UDP-D-glucose + (S)-4-hydroxymandelonitrile = UDP + (S)-4-hydroxymandelonitrile beta-D-glucoside
show the reaction diagram
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-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexosyl group transfer
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of secondary metabolites
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Cyanoamino acid metabolism
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dhurrin biosynthesis
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SYSTEMATIC NAME
IUBMB Comments
UDP-D-glucose:(S)-4-hydroxymandelonitrile beta-D-glucosyltransferase
Acts on a wide range of substrates in vitro, including cyanohydrins, terpenoids, phenolics, hexanol derivatives and plant hormones, in a regiospecific manner [3]. This enzyme is involved in the biosynthesis of the cyanogenic glucoside dhurrin in sorghum, along with EC 1.14.13.41, tyrosine N-monooxygenase and EC 1.14.13.68, 4-hydroxyphenylacetaldehyde oxime monooxygenase. This reaction prevents the disocciation and release of toxic hydrogen cyanide [3].
CAS REGISTRY NUMBER
COMMENTARY hide
55354-52-4
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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TREMBL
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
plant tcd2 mutants deficient in the enzyme have reduced vigor, being dwarfed, with poor root development and low fertility. The mutant plants accumulate numerous dhurrin pathway-derived metabolites, some of which are similar to those observed in transgenic Arabidopsis thaliana expressing the CYP79A1 and CYP71E1 genes. The tcd2 mutant suffers from self-intoxication because Sorghum does not have a feedback mechanism to inhibit the initial steps of dhurrin biosynthesis when the glucosyltransferase activity required to complete the synthesis of dhurrin is lacking. Phenotype, detailed overview
metabolism
physiological function
enzyme UGT85B1 is essential for formation of dhurrin in sorghum with no co-expressed endogenous UDP-glucosyltransferases able to replace it. Presence of metabolites in the tcd2 mutant which are suggested to be derived from dhurrin via endogenous pathways for nitrogen recovery, indicating which enzymes may be involved in such pathways, metabolites identification by LC-MS
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
UDP-alpha-D-glucose + phenol
UDP + phenyl beta-D-glucopyranoside
show the reaction diagram
-
very poor substrate
-
-
?
UDP-D-glucose + (S)-4-hydroxymandelonitrile
UDP + (S)-4-hydroxymandelonitrile beta-D-glucoside
show the reaction diagram
UDP-glucose + (R,S)-mandelonitrile
UDP + prunasin
show the reaction diagram
-
-
-
ir
UDP-glucose + 1-hexanol
UDP + hexyl beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + 2-hydroxy-2-methylbutyronitrile
UDP + ?
show the reaction diagram
-
-
-
-
?
UDP-glucose + 2-hydroxy-3-methoxybenzyl alcohol
UDP + ?
show the reaction diagram
-
-
-
-
?
UDP-glucose + 3-methyl-2-buten-1-ol
UDP + 3-methyl-2-butenyl-beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + 3-methyl-3-buten-1-ol
UDP + 3-methyl-3-butenyl-beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + 4-hydroxybenzaldehyde
UDP + 4-formylphenyl beta-D-glucopyranoside
show the reaction diagram
-
poor substrate, mixture of 4-hydroxybenzaldehyde with NaCN: good substrate
-
-
?
UDP-glucose + 4-hydroxymandelonitrile
UDP + (S)-4-hydroxymandelonitrile beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + acetone cyanohydrin
UDP + ?
show the reaction diagram
-
-
-
-
?
UDP-glucose + alpha-terpinol
UDP + alpha-terpinyl-beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + benzyl alcohol
UDP + benzyl beta-D-glucoside
show the reaction diagram
-
-
-
?
UDP-glucose + beta-citronellol
UDP + beta-citronellol beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + cis-3-hexen-1-ol
UDP + cis-3-hexenyl-beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + farnesol
UDP + farnesyl beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + geraniol
UDP + geranyl-beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + linalool
UDP + linalool beta-D-glucoside
show the reaction diagram
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-
-
-
?
UDP-glucose + mandelonitrile
UDP + 2-(beta-D-glucopyranosyl)-2-phenylacetonitrile
show the reaction diagram
-
-
-
-
?
UDP-glucose + mandelonitrile
UDP + mandelonitrile beta-D-glucoside
show the reaction diagram
specific for UDP-glucose
-
-
?
UDP-glucose + nerol
UDP + nerol beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + trans-2-hexen-1-ol
UDP + trans-2-hexenyl-beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + vanillic acid
UDP + vanillic acid beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDP-glucose + vanillin
UDP + vanillin beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDPalpha-D-glucose + benzoic acid
UDP + 1-O-benzoyl-beta-D-glucose
show the reaction diagram
at 4% of the rate with p-hydroxymandelonitrile
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-
?
UDPglucose + (S)-4-hydroxymandelonitrile
UDP + (S)-4-hydroxymandelonitrile beta-D-glucoside
show the reaction diagram
UDPglucose + (S)-mandelonitrile
UDP + (S)-mandelonitrile beta-D-glucoside
show the reaction diagram
UDPglucose + 4-hydroxybenzoic acid
UDP + glucosyl-4-hydroxybenzoate
show the reaction diagram
-
poor substrate
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-
?
UDPglucose + 4-hydroxybenzyl alcohol
UDP + 4-hydroxybenzyl glucoside
show the reaction diagram
-
at 36% the rate of 4-hydroxymandelonitrile
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-
?
UDPglucose + benzyl alcohol
UDP + benzylglucoside
show the reaction diagram
at 13% of the rate with p-hydroxymandelonitrile
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?
UDPglucose + geraniol
UDP + geraniol glucoside
show the reaction diagram
at 11% of the rate with p-hydroxymandelonitrile
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?
UDPglucose + hydroquinone
UDP + hydroquinone glucoside
show the reaction diagram
-
at 41% the rate of 4-hydroxymandelonitrile
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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
UDP-D-glucose + (S)-4-hydroxymandelonitrile
UDP + (S)-4-hydroxymandelonitrile beta-D-glucoside
show the reaction diagram
UDP-glucose + (R,S)-mandelonitrile
UDP + prunasin
show the reaction diagram
B2XBQ5
-
-
-
ir
UDP-glucose + 4-hydroxymandelonitrile
UDP + (S)-4-hydroxymandelonitrile beta-D-glucoside
show the reaction diagram
-
-
-
-
?
UDPglucose + (S)-4-hydroxymandelonitrile
UDP + (S)-4-hydroxymandelonitrile beta-D-glucoside
show the reaction diagram
additional information
?
-
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final enzyme of dhurrin biosynthesis
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
not activated by Ca2+ or Mg2+
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4-hydroxybenzaldehyde
potent inhibitor, 1 mg/ml bovine serum albumin protects
4-hydroxymandelonitrile
possibly an inhibitor, 1 mg/ml bovine serum albumin protects
benzaldehyde
potent inhibitor, 1 mg/ml bovine serum albumin protects
Dhurrin
potent inhibitor, 1 mg/ml bovine serum albumin protects
mandelonitrile
potent inhibitor, 1 mg/ml bovine serum albumin protects
sambunigrin
potent inhibitor, 1 mg/ml bovine serum albumin protects
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
bovine serum albumin
activates
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dithiothreitol
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requirement, 5 mM
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.66
1-Hexanol
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6.33
2-hydroxy-3-methoxybenzyl alcohol
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10.3
3-methyl-2-buten-1-ol
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0.69
3-methyl-3-buten-1-ol
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0.3
benzyl alcohol
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0.13
beta-citronellol
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0.73
cis-3-hexen-1-ol
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0.14
geraniol
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0.84
mandelonitrile
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1.13
nerol
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0.8
UDP-glucose
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pH 7.5, 30°C, cosubstrate: mandelonitrile
0.029
UDPglucose
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.06
1-Hexanol
Sorghum bicolor
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0.39
2-hydroxy-3-methoxybenzyl alcohol
Sorghum bicolor
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0.91
3-methyl-2-buten-1-ol
Sorghum bicolor
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0.07
3-methyl-3-buten-1-ol
Sorghum bicolor
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0.09
beta-citronellol
Sorghum bicolor
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0.03
cis-3-hexen-1-ol
Sorghum bicolor
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0.1
geraniol
Sorghum bicolor
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10.6
mandelonitrile
Sorghum bicolor
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0.06
nerol
Sorghum bicolor
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8.9
UDP-glucose
Sorghum bicolor
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pH 7.5, 30°C, cosubstrate: mandelonitrile
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8.2 - 8.5
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-
additional information
theoretical pI of 5.3
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
activity is 3fold greater in bitter almonds than in non-bitter types
Manually annotated by BRENDA team
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shoots of etiolated seedlings
Manually annotated by BRENDA team
additional information
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no activity in bundle sheath extracts
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
UGT85B1 is found mainly in the soluble stroma fraction
Manually annotated by BRENDA team
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transient expression of a fluorescent fusion protein in Sorghum bicolor epidermal cells as monitored by confocal laser scanning microscopy shows that UGT85B1-YFP accumulates in the cytoplasm in the absence of CYP79A1 or CYP71E1
Manually annotated by BRENDA team
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transient expression of a fluorescent fusion protein in Sorghum bicolor epidermal cells as monitored by confocal laser scanning microscopy shows that UGT85B1-YFP in the presence of CYP79A1 and CYP71E1 shifts towards the surface of the ER membrane in the periphery of biosynthetic active cells
Manually annotated by BRENDA team
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epidermal protoplasts
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Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
52900
x * 52900, calculated from the amino acid sequence
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 50000-55000, SDS-PAGE; x * 52900, calculated from the amino acid sequence
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
activity is largely uneffected by both freezing at -80°C and addition of glycerol, DTT stabilizes
stable to freezing, DEAE-chromatography decreases stability to freezing
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
0°C, unstable at
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4°C, 2 days, lowering the concentration of DTT from 5 to 2 mM results in a 10fold decrease in activity
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
420fold, native and recombinant enzyme, expressed in Escherichia coli JM109
77fold, partial
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloning of a full-length cDNA encoding enzyme, expression in Escherichia coli JM109, 492-amino acids translation product
expressed in Arabidopsis thaliana
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expressed in Escherichia coli
gene UGT85B1, DNA and amino acid sequence determination and analysis of wild-type and mutant tcd2 genes. The CYP79A1, CYP71E1 and UGT85B1 genes required for dhurrin synthesis are clustered
mutant enzymes, expression in Escherichia coli
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recombinant expression in Nicotiana tabacum chloroplasts, coexpression with CYP79A1 and CYP71E1 linked in a tricistronic operon construct with the essential regulatory elements
transgenic Arabidopsis thaliana plants expressing the entire biosynthetic pathway for the tyrosine-derived cyanogenic glucoside dhurrin as accomplished by insertion of CYP79A1, CYP71E2, and UFT85B1 are shown to accumulate 4% dry-weight dhurrin with marginal inadvertent effects on plant morphology, free amino acid pools, transcriptome, and metabolome
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E410A
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glucosylation of mandelonitrile is 225fold reduced compared to wild-type enzyme
R201A
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glucosylation of mandelonitrile is 20fold reduced compared to wild-type enzyme
S391A
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glucosylation of mandelonitrile is 185fold reduced compared to wild-type enzyme
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
agriculture
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it is possible to engineer plants (Arabidopsis thaliana) which express the high flux pathway for dhurrin synthesis
biotechnology
integration of genes CYP79A1, CYP71E1, and UGT85B1 in Nicotiana tabacum chloroplast genome and functional expression, the enzymes convert endogenous tyrosine into dhurrin using electrons derived directly from the photosynthetic electron transport chain, without the need for the presence of an NADPH-dependent P450 oxidoreductase. The dhurrin produced in the engineered plants amounted to 0.1-0.2% of leaf dry weight compared to 6% in the origin Sorghum bicolor. Plant P450s involved in the synthesis of economically important compounds can be engineered into the thylakoid membrane of chloroplasts, and their full catalytic cycle can be driven directly by photosynthesis-derived electrons
synthesis
integration of genes CYP79A1, CYP71E1, and UGT85B1 in Nicotiana tabacum chloroplast genome and functional expression, the enzymes convert endogenous tyrosine into dhurrin using electrons derived directly from the photosynthetic electron transport chain, without the need for the presence of an NADPH-dependent P450 oxidoreductase. The dhurrin produced in the engineered plants amounted to 0.1-0.2% of leaf dry weight compared to 6% in the origin Sorghum bicolor. Plant P450s involved in the synthesis of economically important compounds can be engineered into the thylakoid membrane of chloroplasts, and their full catalytic cycle can be driven directly by photosynthesis-derived electrons