4.1.2.47: (S)-hydroxynitrile lyase
This is an abbreviated version!
For detailed information about (S)-hydroxynitrile lyase, go to the full flat file.
Word Map on EC 4.1.2.47
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4.1.2.47
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esculenta
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manihot
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brasiliensis
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hevea
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synthesis
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cassava
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ketone
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hcn
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lyases
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benzaldehyde
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s-mandelonitrile
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rubber
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athnl
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crantz
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hydrocyanic
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s-cyanohydrins
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r-selective
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arylacetonitrilase
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s-enantiomer
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cyanogenesis
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montanum
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s-mandelic
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pharmacology
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agriculture
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analysis
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drug development
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food industry
- 4.1.2.47
- esculenta
- manihot
- brasiliensis
- hevea
- synthesis
- cassava
- ketone
- hcn
- lyases
- benzaldehyde
-
s-mandelonitrile
- rubber
- athnl
- crantz
-
hydrocyanic
-
s-cyanohydrins
-
r-selective
- arylacetonitrilase
-
s-enantiomer
-
cyanogenesis
- montanum
-
s-mandelic
- pharmacology
- agriculture
- analysis
- drug development
- food industry
Reaction
Synonyms
(R)-LuHNL, (R)-Oxynitrilase, (S)-cyanohydrin producing hydroxynitrile lyase, (S)-Hb-HNL, (S)-HbHNL, (S)-HNL, (S)-Hydroxynitrile lyase, (S)-Me-HNL, (S)-MeHNL, (S)-Oxynitrilase, (S)-selective HNL, (S)-selective hydroxynitrile lyase, Acetone cyanohydrin lyase, ACL, alpha-Hydroxynitrile lyase, EC 4.1.2.37, EC 4.1.2.39, Hb-HNL, HbHNL, HNL, HNL1, Hydroxynitrile lyase, LuHNL, MeHNKL, MeHNL, mut-HNL1, Oxynitrilase, S-HnL, S-hydroxynitrile lyase, S-selective HnL, S-selective hydroxynitrile lyase, S-stereoselective HNL
ECTree
4.1.2.47 (S)-hydroxynitrile lyaseAdvanced search results
results (
results (Engineering
Engineering on EC 4.1.2.47 - (S)-hydroxynitrile lyase
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PROTEIN VARIANTS 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
H103C
the mutated enzyme shows low enantioselectivity and specific activity for (S)-mandelonitrile synthesis. (S)-Mandelonitrile enantiomeric excess is 60%
H103C/N156D
hydroxynitrile lyase of mutant enzyme H103C/N156D is approximately four times higher than that for mutant enzyme H103C. (S)-Mandelonitrile enantiomeric excess is 32%
H103C/N156G
C81L
C81S
E79A
F125T
F125T/L146M
the mutant shows higher specific activity towards racemic mandelonitrile compared to the wild type enzyme
F125T/Y133F
the mutant shows higher specific activity towards racemic mandelonitrile compared to the wild type enzyme
F210I
H103L/W128A
increased activity with the substrate (2S)-hydroxy[4-(methoxy)cyclohex-3-en-1-yl]ethanenitrile compared to the starting clone W128A
H10A
mutation results in a 30000 Da protein with increased electrophoretic mobility on native high percentage (16%) polyacrylamide gels. the mutant enzyme displays almost wild-type specific activity in crude extracts, suggesting that His10 is not crucial for activity. However, activity is almost completely lost during purification, supporting the possibility that the H10A exchange has a destabilizing effect and may prevent formation of an active dimer of the enzyme after purification
H235A
I12A
mutant yields mostly insoluble protein, which suggests that the substitution hinders protein folding
I209A
I209G
K236L
inactive mutant protein, three-dimensional structure is similar to wild-type enzyme
L121F
6.9fold increase in specificity constants (kcat/Km) for racemic mandelonitrile
L121Y
L121Y/F125T
L121Y/F125T/L146M
L121Y/L146M
the mutant shows higher specific activity towards racemic mandelonitrile compared to the wild type enzyme
L146M
the mutant shows lower specific activity towards racemic mandelonitrile compared to the wild type enzyme
L148F
the mutant shows lower specific activity towards racemic mandelonitrile compared to the wild type enzyme
L152F
S80A
V106F
the mutant shows lower specific activity towards racemic mandelonitrile compared to the wild type enzyme
V106F/L121Y
the mutant shows higher specific activity towards racemic mandelonitrile compared to the wild type enzyme
V106F/L121Y/F125T
W128A
W128A/I219V
very weak activity in cleavage reaction with (2S)-hydroxy[4-(methoxy)cyclohex-3-en-1-yl]ethanenitrile
W128A/K147R
very weak activity in cleavage reaction with (2S)-hydroxy[4-(methoxy)cyclohex-3-en-1-yl]ethanenitrile
W128A/P187L
no activity in cleavage reaction with (2S)-hydroxy[4-(methoxy)cyclohex-3-en-1-yl]ethanenitrile
W128A/Q215H
slightly increased activity with the substrate (2S)-hydroxy[4-(methoxy)cyclohex-3-en-1-yl]ethanenitrile compared to the starting clone W128A
Y133F
the mutant shows lower specific activity towards racemic mandelonitrile compared to the wild type enzyme
C81A
D208A
Km-value for 2-hydroxy-2-methylpropanenitrile increases from 101 mM for wild-type enzyme to over 200 mM for the mutant enzyme
G113S
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enhanced thermal stability compared to wild-type enzyme, mutant enzyme retains slight higher activity than the wild-type enzyme in an acidic environment, so the mutant enzyme maybe more effective for synthesis of (S)-cyanohydrin than the wild-type enzyme
H103C
the mutant displays 9.3fold increase in total specific activity in the cell-free extract compared with the wild type
H103I
the mutant displays 8.1fold increase in total specific activity in the cell-free extract compared with the wild type
H103L
H103M
the mutant displays 9.07fold increase in total specific activity in the cell-free extract compared with the wild type
H103Q
the mutant displays 4.06fold increase in total specific activity in the cell-free extract compared with the wild type
H103S
the mutant displays 2.9fold increase in total specific activity in the cell-free extract compared with the wild type
H103T
the mutant displays 3.4fold increase in total specific activity in the cell-free extract compared with the wild type
H236A
mutant enzyme is unable to catalyze the decomposition of 2-hydroxy-2-methylpropanenitrile
K176P
the mutant displays 2.02fold increase in total specific activity in the cell-free extract compared with the wild type
K176P/K199P/K224P
the mutant displays 6.97fold increase in total specific activity in the cell-free extract compared with the wild type
K176P/K224P
the mutant displays 5.05fold increase in total specific activity in the cell-free extract compared with the wild type
K199P
the mutant displays 1.38fold increase in total specific activity in the cell-free extract compared with the wild type
K199P/K224P
the mutant displays 4.25fold increase in total specific activity in the cell-free extract compared with the wild type
K224P
the mutant displays 2.53fold increase in total specific activity in the cell-free extract compared with the wild type
S80A
mutant enzyme is completely inactive in the 2-hydroxy-2-methylpropanenitrile cleaving assay. No differences to wild type MeHNL according to oligomeric structure, molecular weight, and behavior in the standard purification procedure
W128A
W128C
W128L
W128Y
additional information
H103C/N156G
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the double mutant is suitable for (S)-mandelonitrile synthesis, improving its enantioselectivity and specific activity. The specific activity of the H103C/N156G mutant enzyme is improved to 154 U/mg from 52 U/mg as compared with wild type enzyme for (S)-mandelonitrile production, and the enantiomeric excess of (S)-mandelonitrile produced by the double mutant is increased to 93% from 55% compared with wild type enzyme
H103C/N156G
the specific activity of the H103C/N156G mutant for (S)-mandelonitrile production is raised to 154 U/mg (wild-type hydroxynitrile lyase: 52 U/mg). The enantiomeric excess is increased to 93% (wild-type: 55%). Km-value for (R)-mandelonitrile and kcat for (S)-mandelonitrile increase by the mutation at Asn156, thus contributing to the increase in enantiomeric excess
C81L
the mutant has high enantioselectivity towards (R)-mandelonitrile with a kcat value 1.3times higher compared to the wild type enzyme
C81S
no difference can be observed in the electrophoretic mobilities between the wild-type and mutant protein on native polyacrylamide gels and by isoelectric focusing. Mutation does not change the charge or size of the amino acid side chain, but nevertheless greatly reduces activity
E79A
no difference can be observed in the electrophoretic mobilities between the wild-type and mutant protein on native polyacrylamide gels and by isoelectric focusing. Mutation greatly reduces, but does not abolish activity. The negative charge provided by Glu-79 may be required in the active site, but a direct participation of this residue in enzyme catalysis is not suggested
F125T
2.6fold increase in kcat for racemic mandelonitrile, 4.5fold increase in specificity constants (kcat/Km) for racemic mandelonitrile
F125T
the mutant has high enantioselectivity towards (R)-mandelonitrile with a kcat value 2.6times and a kcat/Km value 4.5times higher compared to the wild type enzyme
F210I
mutant yields mostly insoluble protein, which suggests that the substitution hinders protein folding
F210I
the mutant shows lower specific activity towards racemic mandelonitrile compared to the wild type enzyme
H235A
inactive mutant enzyme, no difference can be observed in the electrophoretic mobilities between the wild-type and mutant protein on native polyacrylamide gels and by isoelectric focusing
I209A
3.9fold increase in specificity constants (kcat/Km) for racemic mandelonitrile
I209A
the mutant has higher enantioselectivity towards (R)-mandelonitrile with a kcat/Km value 3.9times higher compared to the wild type enzyme
I209G
mutant yields mostly insoluble protein, which suggests that the substitution hinders protein folding, moderate substrate inhibition by rac-mandelonitrile
I209G
the mutant shows lower specific activity towards racemic mandelonitrile compared to the wild type enzyme
L121Y
5.3fold increase in specificity constants (kcat/Km) for racemic mandelonitrile, 4.2fold increase in kcat for racemic mandelonitrile, the mutant enzyme shows the same high (S)-enantioselectivity (98% ee) as wild-type enzyme. The mutant catalyzes the cleavage of 2-nitro-1-phenylethanol 4.8 times more efficiently than the wild type enzyme, this is similar to the 4.5-fold increase seen for mandelonitrile cleavage
L121Y
the mutant has high enantioselectivity towards (S)-mandelonitrile with a kcat value 4.2times higher and a kcat/Km value 5.3times higher compared to the wild type enzyme
L121Y/F125T
1.7fold increase in kcat for racemic mandelonitrile, 1.9fold increase in specificity constants (kcat/Km) for racemic mandelonitrile
L121Y/F125T
the mutant has high enantioselectivity towards (R)-mandelonitrile with a kcat value 4.2times higher and a kcat/Km value 1.7times higher compared to the wild type enzyme
L121Y/F125T
the mutant shows higher specific activity towards racemic mandelonitrile compared to the wild type enzyme
L121Y/F125T/L146M
5.5fold increase in activity over the wild type enzyme
L121Y/F125T/L146M
the mutant shows higher specific activity towards racemic mandelonitrile compared to the wild type enzyme
L152F
1.5fold increase in specificity constants (kcat/Km) for racemic mandelonitrile
L152F
the mutant has higher enantioselectivity towards (R)-mandelonitrile with a kcat/Km value 1.5times higher compared to the wild type enzyme
S80A
no difference can be observed in the electrophoretic mobilities between the wild-type and mutant protein on native polyacrylamide gels and by isoelectric focusing
V106F/L121Y/F125T
3.3fold increase in kcat for racemic mandelonitrile, 5.5fold increase in specificity constants (kcat/Km) for racemic mandelonitrile
V106F/L121Y/F125T
the mutant has high enantioselectivity towards (R)-mandelonitrile with a kcat value 3.3times higher compared to the wild type enzyme
W128A
higher conversion and better selectivity than wild-type enzyme with the substrate 4-methoxycyclohex-3-ene-1-carbaldehyde
W128A
lower conversion and lower selectivity than wild-type enzyme with the substrate 4-methoxycyclohex-3-ene-1-carbaldehyde
H103L
the mutant displays 11.1fold increase in total specific activity in the cell-free extract compared with the wild type
W128A
activity with the natural substrate 2-hydroxy-2-methylpropanenitrile (acetone cyanohydrin) is 70% of wild-type activity. The specific activities of MeHNL-W128A for the unnatural substrates mandelonitrile and 4-hydroxymandelonitrile are increased 9fold and 450fold, respectively, compared with the wild-type. The crystal structure of the mutant W128A substrate free form at 2.1 A resolution indicates that the W128A substitution has significantly enlarged the active-site channel entrance, and thereby explains the observed changes in substrate specificity for bulky substrates
W128A
although the variant W128A shows a higher activity with respect to (S)-3-phenoxy-benzaldehyde cyanohydrin, the enantioselectivity is reduced to 85%, compared to 97% of wild-type enzyme
W128A
substitution of tryptophan128 by an alanine residue enlarges the entrance channel to the active site of MeHNL and thus facilitates access of sterically demanding substrates to the active site, increased conversion rate towards 3-phenoxybenzaldehyde, octan-3-one and heptan-3-one
W128C
mutation increases the specific activity towards 4-hydroxymandelonitrile of the various MeHNL mutant compared to the wild-type enzyme
W128L
mutation increases the specific activity towards 4-hydroxymandelonitrile of the various MeHNL mutant compared to the wild-type enzyme
W128Y
mutation increases the specific activity towards 4-hydroxymandelonitrile of the various MeHNL mutant compared to the wild-type enzyme
additional information
C-terminally truncated enzymes exhibit activities of 75, 102, 65 and 48 U/l for enzyme after removing 2, 4, 6, and 8 amino acids, respectively
additional information
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C-terminally truncated enzymes exhibit activities of 75, 102, 65 and 48 U/l for enzyme after removing 2, 4, 6, and 8 amino acids, respectively
