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Information on EC 5.1.2.2 - mandelate racemase and Organism(s) Pseudomonas putida and UniProt Accession P11444
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5.1.2.2 mandelate racemaseSpecify your search results
Word Map
- 5.1.2.2
- s-mandelate
-
kenyon
-
alpha-proton
- muconate
-
gerlt
-
1,1-proton
- benzohydroxamate
-
petsko
-
kozarich
- mitra
- galactarate
- benzoylformate
- d-glucarate
-
ransom
- synthesis
- analysis
- biotechnology
The taxonomic range for the selected organisms is: Pseudomonas putida
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota
Synonyms
mandelate racemase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
mandelic acid racemase
-
-
-
-
MR
racemase, mandelate
-
-
-
-
MR
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
(S)-mandelate = (R)-mandelate
His297 and Asp270 function together as a catalytic dyad
(S)-mandelate = (R)-mandelate
the enzyme utilizes a two-base mechanism with Lys166 and His297 acting as Broensted acid and base catalysts, respectively, in the R -> S reaction direction. In the S -> R reaction direction, their roles are reversed
(S)-mandelate = (R)-mandelate
enzyme does not catalyze an intermolecular proton transfer to achieve racemization. The enzyme operates via a one-acceptor mechanism, in which the proton abstracted from one stereochemical face of a substrate molecule is returned to the opposite face of the same carbon of the substrate molecule
-
(S)-mandelate = (R)-mandelate
reaction proceeds via a two-base mechanism in which Lys166 abstracts the alpha-proton from (S)-mandelate and His297 abstracts the alpha-proton from (R)-mandelate
-
(S)-mandelate = (R)-mandelate
pathway involving a carbanion intermediate
-
(S)-mandelate = (R)-mandelate
two-base mechanism in which the conjugate acid of the R-specific base is monoprotic and in which the S-specific base may be an amino group
-
(S)-mandelate = (R)-mandelate
the enzyme contains two distinct general acid/base catalysts: Lys166 which abstracts the alpha-proton from (S)-mandelate, and His297, which abstracts the alpha-proton from (R)-mandelate
-
(S)-mandelate = (R)-mandelate
one-base acceptor mechanism, functional asymmetry at the active site
-
(S)-mandelate = (R)-mandelate
one pot enantioconvergent synthesis of R-mandelic acid ester from racemic mandelic acid is achieved by a novel aqueous/organic two-phase system with two-enzyme process consisting of (1) lipase-catalyzed enantioselective esterification of racemic mandelic acid in organic solvent and (2) in situ racemization of unreacted isomer, S-mandelic acid in aqueous phase by recombinant mandelate racemase
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
racemization
-
-
-
-
PATHWAY SOURCE
PATHWAYS
-
-, -
SYSTEMATIC NAME
IUBMB Comments
mandelate racemase
-
CAS REGISTRY NUMBER
COMMENTARY
9024-04-8
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
(2R)-2-naphthylglycolate
(2S)-2-naphthylglycolate
a non-natural substrate
-
-
?
(R)-3-chloromandelate
(S)-3-chloromandelate
a non-natural substrate
-
-
?
(S)-2-chloromandelate
(R)-2-chloromandelate
binding structure
-
-
r
(S)-vinylglycolate
(R)-vinylglycolate
two-step quite symmetric process through a dianionic enolic intermediate that is formed after the abstraction of the alpha-protein of vinylglycolate by a basic enzymatic residue and is then reprotonated by another residue
-
?
(2R)-2-(4-bromophenyl)-2-hydroxyacetamide
?
-
22% of the activity with (R)-mandelate
-
-
?
(2R)-2-furyl(hydroxy)acetic acid
?
-
24% of the activity with (R)-mandelate
-
-
?
(2R)-2-hydroxy-2-phenylacetamide
?
-
15% of the activity with (R)-mandelate
-
-
?
(2R)-2-hydroxy-3-(1-H-imidazol-1-yl)propanoic acid
?
-
5.4% of the activity with (R)-mandelate
-
-
?
(2R)-2-hydroxypentanoic acid
?
-
1% of the activity with (R)-mandelate
-
-
?
(2R)-3-furyl(hydroxy)acetic acid
?
-
38% of the activity with (R)-mandelate
-
-
?
(2R)-cyclohex-1-en-1-yl(hydroxy)acetic acid
?
-
50% of the activity with (R)-mandelate
-
-
?
(2R)-hydroxy(2-naphthyl)acetic acid
?
-
26% of the activity with (R)-mandelate
-
-
?
(2S)-hydroxy(2-thienyl)acetic acid
?
-
59% of the activity with (R)-mandelate
-
-
?
(R)-(E)-2-hydroxy-3-pentenoic acid
?
-
36% of the activity with (R)-mandelate
-
-
?
(R)-(E)-2-hydroxy-4-phenyl-3-butenoic acid
?
-
53% of the activity with (R)-mandelate
-
-
?
(S)-mandelic acid amide
(R)-mandelic acid amide
-
activity is enhanced by an electron-withdrawing substituent in the phenyl moiety
-
?
2-hydroxybut-3-enoic acid
?
-
35% of the activity with (R)-mandelate
-
-
?
4-bromo-(R)-mandelate
4-bromo-(S)-mandelate
-
376% of the activity with (R)-mandelate
-
-
r
4-chloro-(R)-mandelate
4-chloro-(S)-mandelate
-
326% of the activity with (R)-mandelate
-
-
r
4-fluoro-(R)-mandelate
4-fluoro-(S)-mandelate
-
96% of the activity with (R)-mandelate
-
-
r
4-hydroxy-(R)-mandelate
4-hydroxy-(S)-mandelate
-
45% of the activity with (R)-mandelate
-
-
r
4-methoxy-(R)-mandelate
4-methoxy-(S)-mandelate
-
17% of the activity with (R)-mandelate
-
-
r
5-chloro-(R)-mandelate
5-chloro-(S)-mandelate
-
61% of the activity with (R)-mandelate
-
-
r
(R)-mandelate
(S)-mandelate
wild-type enzyme shows slightly higher affinity for (S)-mandelate than for (R)-mandelate, but catalyzes the turnover of (R)-mandelate slightly more rapidly
-
-
r
(S)-mandelate
(R)-mandelate
catalyzes the Mg2+-dependent 1,1-proton transfer that interconverts the enantiomers, catalysis is done via a two-base mechanism
-
-
r
(S)-2-hydroxy-3-butenoic acid
(R)-2-hydroxy-3-butenoic acid
-
maximal racemization rate is 35% relative to mandelate
-
?
D-mandelate
L-mandelate
-
strictly inducible enzyme. Inducers: D-mandelate, L-mandelate, benzoylformate
-
-
?
D-mandelate
L-mandelate
-
first enzyme in the bacterial pathway that converts mandelic acid to benzoic acid
-
-
?
D-mandelate
L-mandelate
-
first enzyme of pathway of oxidation of D,L-mandelate that permits certain strains of Pseudomonas putida to use mandelate as a sole source of carbon and energy
-
-
?
p-(bromomethyl)mandelate
p-(methyl)benzoylformate + Br-
-
-
i.e. alpha-carboxy-alpha-hydroxy-p-xylylene
?
additional information
?
-
mandelate racemase catalyzes the Mg2+-dependent 1,1-proton transfer that interconverts the enantiomers of mandelate
-
-
?
additional information
?
-
-
mandelate racemase catalyzes the Mg2+-dependent 1,1-proton transfer that interconverts the enantiomers of mandelate
-
-
?
additional information
?
-
substrate recognition and binding method, overview. Mandelate racemase from Pseudomonas putida catalyzes the interconversion of the enantiomers of mandelic acid and a variety of aryl- and heteroaryl-substituted mandelate derivatives. beta,gamma-Unsaturation is not an absolute requirement for catalysis and that mandelate racemase can bind and catalyze the racemization of (S)- and (R)-trifluorolactate. The enzyme catalyzes hydrogen-deuterium exchange at the alpha-postion of trifluorolactate
-
-
?
additional information
?
-
-
substrate recognition and binding method, overview. Mandelate racemase from Pseudomonas putida catalyzes the interconversion of the enantiomers of mandelic acid and a variety of aryl- and heteroaryl-substituted mandelate derivatives. beta,gamma-Unsaturation is not an absolute requirement for catalysis and that mandelate racemase can bind and catalyze the racemization of (S)- and (R)-trifluorolactate. The enzyme catalyzes hydrogen-deuterium exchange at the alpha-postion of trifluorolactate
-
-
?
additional information
?
-
enzyme activity detection in the high-throughput screening method of enzymes involving the racemization of (S)-2-chloromandelate, method evaluation, overview
-
-
?
additional information
?
-
-
not racemized: 4-hydroxy-3-methoxymandelate, 3,4-dihydroxymandelate
-
-
?
additional information
?
-
-
efficient racemization of 2-hydroxy-2-heteroaryl-acetic acid derivatives. Steric limitations for aryl-substituted mandelate derivatives are elucidated to be particularly striking for o-substituents, whereas m- and p-analogues are freely accepted as well as heteroaryl and naphthyl analogs. The electronic character of substituents is found to play an important role: whereas electron-withdrawing substituents dramatically enhance the racemization rates, electron-donating analogs cause a depletion
-
?
additional information
?
-
-
no activity with phenyl glycine and mandelic acid hydrazide
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
(S)-mandelate
(R)-mandelate
catalyzes the Mg2+-dependent 1,1-proton transfer that interconverts the enantiomers, catalysis is done via a two-base mechanism
-
-
r
D-mandelate
L-mandelate
-
strictly inducible enzyme. Inducers: D-mandelate, L-mandelate, benzoylformate
-
-
?
D-mandelate
L-mandelate
-
first enzyme in the bacterial pathway that converts mandelic acid to benzoic acid
-
-
?
D-mandelate
L-mandelate
-
first enzyme of pathway of oxidation of D,L-mandelate that permits certain strains of Pseudomonas putida to use mandelate as a sole source of carbon and energy
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mg2+
Co2+
Divalent metal ions
Fe2+
-
absolute requirement for a divalent metal ion: Mg2+, Mn2+, Ni2+, Co2+ or Fe2+
Mg2+
Mn2+
Ni2+
Co2+
-
Mg2+, Mn2+, Ni2+, Co2+ or Fe2+ satisfy the divalent metal ion requirement
Divalent metal ions
-
increase the affinity of the enzyme for the substrate. No absolute requirement
Divalent metal ions
-
the required divalent metal ion is ligated by the carboxyl groups of Asp195, Glu221, and Glu247, which emanate from the carboxy-terminal ends of strands 3, 4 and 5, respectively
Mg2+
-
Mg2+, Mn2+, Ni2+, Co2+ or Fe2+ satisfy the divalent metal ion requirement
Mg2+
-
Km: 0.65 mM for wild-type enzyme and 0.34 mM for mutant enzyme N197A
Mn2+
-
Mg2+, Mn2+, Ni2+, Co2+ or Fe2+ satisfy the divalent metal ion requirement
Mn2+
-
enzyme binds 0.9 mol Mn2+ per mol of subunit with a dissociation constant of 0.008 mM. Also six additional Mn2+ bind to the enzyme much more weakly, with a dissociation constant of 1.5 mM
Ni2+
-
Mg2+, Mn2+, Ni2+, Co2+ or Fe2+ satisfy the divalent metal ion requirement
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
(R,S)-1-hydroxyethylphosphonate
transition state analogue inhibitor
(R,S)-2,2,2-trifluoro-1-hydroxyethylphosphonate
transition state analogue inhibitor
(R,S)-alpha-hydroxybenzylphosphonate
transition state analogue inhibitor
3,3,3-trifluoro-2-hydroxy-2-(trifluoromethyl)-propanoate
a substrate-product analogue and a potent competitive inhibitor with both (R)-mandelate and (R)-trifluorolactate. The inhibitor exhibits a different binding mode with the two trifluoromethyl groups closely packed against the 20s loop and the carboxylate bridging the two active site Broensted acid-base catalysts Lys166 and His297
3-hydroxypyruvate
an irreversible, time-dependent inhibitor, causes inactivation of mandelate racemase. Protection from inactivation by the competitive inhibitor benzohydroxamate. 3-Hydroxypyruvate undergoes Schiff-base formation with Lys166 at the active site, followed by formation of an aldehyde/enol(ate) adduct
Cupferron
a reasonable mimic of the transition state and/or intermediate that chelates the active site divalent metal ion and is bound in a conformation with the phenyl ring coplanar with the diazeniumdiolate moiety, active site binding, structure comparison with bound benzohydroxamate, overview
N-hydroxyformanilide
-
potent competitive inhibitor, MR can bind either the protonated or deprotonated forms of N-hydroxyformanilide, with a 10fold greater affinity for the latter form
benzohydroxamate
a reasonable mimic of the transition state and/or intermediate that chelates the active site divalent metal ion and is bound in a conformation with the phenyl ring coplanar with the hydroxamate moiety, active site binding, structure comparison with bound Cupferron, overview
benzohydroxamate
BzH, the enzyme binds the intermediate/transition state analogue inhibitor in an entropy-driven process, consistent with an increased number of hydrophobic interactions, additional specific interactions contribute to binding, detailed kinetics and binding analysis, overview
DL-alpha-Phenylglycidate
-
binds at the active site of the enzyme, complete and irreversible inhibition in presence of Mg2+. Both D- and L-mandelate protect from inactivation
EDTA
-
activity is fully restored upon addition of certain divalent metal ions. Mg2+ is the most effective, followed by Co2+, Ni2+, Mn2+ and Fe2+
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
sucrose
higher viscosity by addition of up to 35% sucrose elevates the enzyme activity
sucrose
-
slight activating effect of sucrose on mutant enzyme efficiency. In presence of polymeric viscosogens poly(ethylene glycol) and Ficoll, no effect on turnover number or the ratio of turnover number and Km-value for the wild-type enzyme is observed
additional information
-
activation of mandelate racemase via immobilization in lyotropic liquid crystals for biocatalysis in organic solvents: application and modeling
-
additional information
-
the activity of the enzyme immobilized through ionic binding onto DEAE- and TEAE-cellulose is significantly enhanced as compared to the native protein, i.e. 2.7fold and 2.5fold
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
1.2
(R)-trifluorolactate
pH 7.5, 25°C, recombinant wild-type enzyme
1.74
(S)-trifluorolactate
pH 7.5, 25°C, recombinant wild-type enzyme
3.5
(2R)-2-naphthylglycolate
pH 7.5, 25°C, wild-type enzyme, in presence of 20 mM MgCl2
3.5
(R)-2-naphthylglycolate
wild-type, pH 7.5, 25°C, presence of 20 mM Mg2+
3.7
(R)-2-naphthylglycolate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
4.2
(R)-2-naphthylglycolate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
1.1
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V29F
-
1.2
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, wild-type
-
1.3
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26L
-
1.6
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V29L
-
1.7
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26F
-
1.8
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V29A
-
1.9
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant A25V
-
1.9
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V22A
-
2.1
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26A
-
2.2
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V22F
-
3.2
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant T24S
-
3.7
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V22I/V29L
-
4.2
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V22I
-
4.4
(R)-2-naphtylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26A/V29L
-
1.2
(R)-mandelate
pH 7.5, 25°C, recombinant wild-type enzyme
2.8
(R)-mandelate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
2.9
(R)-mandelate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
0.55
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V29F
0.62
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V22F
0.62
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26L
0.74
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26A
0.8
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V29L
0.93
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V29A
1.1
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26F
1.4
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant T24S
1.55
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V22A
1.6
(S)-2-naphthylglycolate
wild-type, pH 7.5, 25°C, presence of 20 mM Mg2+
1.67
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant A25V
1.7
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26A/V29L
2.4
(S)-2-naphthylglycolate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
2.4
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V26A/V29L
2.9
(S)-2-naphthylglycolate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
2.9
(S)-2-naphthylglycolate
pH 7.5 at 25°C, 3.3 mM Mg2+, mutant V22I
0.3
(S)-Mandelate
recombinant mandelate racemase, putative misfolded form with N-terminal hexahistidine tag, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
0.6
(S)-Mandelate
recombinant mrIII fusion protein bearing an N-terminal StrepII-tag and a C-terminal decahistidine tag, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
0.8
(S)-Mandelate
recombinant mandelate racemase represents correctly folded enzyme with N-terminal hexahistidine tag, in Na-HEPES buffer (0.1 M, pH 7.5),3.3 mM MgCl2
1.1
(S)-Mandelate
recombinant mandelate racemase, replacement of the N-terminal hexahistidine tag by a StrepII-tag appears to ameliorate the folding problem yielding a single enzyme species, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
1.5
(S)-Mandelate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
1.7
(S)-Mandelate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
0.04
Mg2+
pH 7.5, mutant V22A, addition of either 10 mM (R)- or (S)-mandelate
0.04
Mg2+
pH 7.5, mutant V22F, addition of either 10 mM (R)- or (S)-mandelate
0.05
Mg2+
pH 7.5, mutant V26F, addition of either 10 mM (R)- or (S)-mandelate
0.08
Mg2+
pH 7.5, mutant V29A, addition of either 10 mM (R)- or (S)-mandelate
0.12
Mg2+
pH 7.5, mutant V26L, addition of either 10 mM (R)- or (S)-mandelate
0.15
Mg2+
pH 7.5, mutant V26A, addition of either 10 mM (R)- or (S)-mandelate
0.22
Mg2+
pH 7.5, mutant V29F, addition of either 10 mM (R)- or (S)-mandelate
0.29
Mg2+
pH 7.5, mutant T24S, addition of either 10 mM (R)- or (S)-mandelate
0.34
Mg2+
pH 7.5, wild-type, addition of either 10 mM (R)- or (S)-mandelate
0.47
Mg2+
pH 7.5, mutant V29L, addition of either 10 mM (R)- or (S)-mandelate
0.51
Mg2+
pH 7.5, mutant V26A/V29L, addition of either 10 mM (R)- or (S)-mandelate
0.64
Mg2+
pH 7.5, mutant A25V, addition of either 10 mM (R)- or (S)-mandelate
3.8
Mg2+
pH 7.5, mutant V22I, addition of either 10 mM (R)- or (S)-mandelate
4.2
Mg2+
pH 7.5, mutant V22I/V29L, addition of either 10 mM (R)- or (S)-mandelate
additional information
additional information
Michaelis-Menten kinetics
-
additional information
additional information
-
Michaelis-Menten kinetics
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
2.5
(S)-trifluorolactate
pH 7.5, 25°C, recombinant wild-type enzyme
70
(2R)-2-naphthylglycolate
pH 7.5, 25°C, wild-type enzyme, in presence of 20 mM MgCl2
10.5
(R)-2-naphthylglycolate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
70
(R)-2-naphthylglycolate
wild-type, pH 7.5, 25°C, presence of 20 mM Mg2+
147
(R)-2-naphthylglycolate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
10.5
(R)-2-naphtylglycolate
pH 7.5, 3.3 mM Mg2+, mutant V22I/V29L
-
1010
(R)-mandelate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
1150
(R)-mandelate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
7.1
(S)-2-naphthylglycolate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
38.4
(S)-2-naphthylglycolate
wild-type, pH 7.5, 25°C, presence of 20 mM Mg2+
101
(S)-2-naphthylglycolate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
190
(S)-Mandelate
recombinant mandelate racemase, putative misfolded form with N-terminal hexahistidine tag, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
472
(S)-Mandelate
recombinant mandelate racemase fusion protein bearing an N-terminal StrepII-tag and a C-terminal decahistidine tag, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
710
(S)-Mandelate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
740
(S)-Mandelate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
940
(S)-Mandelate
recombinant mandelate racemase represents correctly folded enzyme, with N-terminal hexahistidine tag, in Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
1124
(S)-Mandelate
recombinant enzyme replacement of the N-terminal hexahistidine tag by a StrepII-tag appears to ameliorate the folding problem yielding a single enzyme species, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
additional information
additional information
dependence of turnover number on pH
-
additional information
additional information
-
dependence of turnover number on pH
-
additional information
additional information
-
dependence of turnover number on pH
-
additional information
additional information
-
the turnover number of the mutant enzyme is unaffected by varying solvent viscosity
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
1.6
(R)-trifluorolactate
pH 7.5, 25°C, recombinant wild-type enzyme
1.4
(S)-trifluorolactate
pH 7.5, 25°C, recombinant wild-type enzyme
20.1
(2R)-2-naphthylglycolate
pH 7.5, 25°C, wild-type enzyme, in presence of 20 mM MgCl2
1100
(S)-Mandelate
recombinant mandelate racemase, replacement of the N-terminal hexahistidine tag by a StrepII-tag appears to ameliorate the folding problem yielding a single enzyme species, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
1200
(S)-Mandelate
recombinant mandelate racemase, represents correctly folded enzyme with N-terminal hexahistidine tag, in Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
7300
(S)-Mandelate
recombinant mandelate racemase, putative misfolded form with N-terminal hexahistidine tag, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
8000
(S)-Mandelate
recombinant mandelate racemase, fusion protein bearing an N-terminal StrepII-tag and a C-terminal decahistidine tag, Na-HEPES buffer (0.1 M, pH 7.5), 3.3 mM MgCl2
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
40.2
(R,S)-1-hydroxyethylphosphonate
pH 7.5, 25°C, recombinant wild-type enzyme
0.67
(R,S)-2,2,2-trifluoro-1-hydroxyethylphosphonate
pH 7.5, 25°C, recombinant wild-type enzyme
0.0047
(R,S)-alpha-hydroxybenzylphosphonate
pH 7.5, 25°C, recombinant wild-type enzyme
0.027
3,3,3-trifluoro-2-hydroxy-2-(trifluoromethyl)-propanoate
pH 7.5, 25°C, recombinant wild-type enzyme
1.3
3-Fluoropyruvate
pH and temperature not specified in the publication
5.5
alpha-hydroxyisobutyrate
pH 7.5, 25°C, recombinant wild-type enzyme
0.023
2-naphtholhydroxamate
wild-type, pH 7.5, 25°C, presence of 20 mM Mg2+
0.112
2-naphtholhydroxamate
mutant V22I/V29L, pH 7.5, 25°C, presence of 20 mM Mg2+
0.19
2-naphtholhydroxamate
mutant V22I, pH 7.5, 25°C, presence of 20 mM Mg2+
0.0047
(R,S)-alpha-hydroxybenzylphosphonate
-
25°C, pH 7.5, wild-type enzyme
0.238
(R,S)-alpha-hydroxybenzylphosphonate
-
25°C, pH 7.5, mutant enzyme N197A
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.00057
wild-type enzyme, pH 7.5, 25°C, substrate (S)-2-chloromandelate
0.00065
wild-type enzyme, pH 7.5, 25°C, substrate (R)-2-chloromandelate
0.00134
mutant V29I/Y45F, pH 7.5, 25°C, substrate (S)-2-chloromandelate
0.00143
0.00149
mutant V22I/V29I, pH 7.5, 25°C, substrate (S)-2-chloromandelate
0.00152
mutant V29I/Y45F, pH 7.5, 25°C, substrate (R)-2-chloromandelate
0.00155
mutant V22I/V29I, pH 7.5, 25°C, substrate (R)-2-chloromandelate
0.00174
mutant V22I/V29I/Y54F, pH 7.5, 25°C, substrate (R)-2-chloromandelate
0.00175
mutant V22I/Y45F, pH 7.5, 25°C, substrate (S)-2-chloromandelate
0.00194
mutant V22I/V29I/Y54F, pH 7.5, 25°C, substrate (S)-2-chloromandelate
additional information
0.00143
mutant V22I/Y45F, pH 7.5, 25°C, substrate (R)-2-chloromandelate
additional information
-
applicability of the electrochemical quarz crystal nanobalance technique to measure the adsorption behavior of an enzyme and its substrate at a platinum electrode
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.5
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
25
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
mandelate racemase is a member of the enolase superfamily. The ability of the enzyme to form and deprotonate a Schiff-base intermediate furnishes a mechanistic link to other alpha/beta-barrel enzymes utilizing Schiff-base chemistry and is in accord with the sequence- and structure-based hypothesis that members of the metal-dependent enolase superfamily and the Schiff-base-forming N-acetylneuraminate lyase superfamily and aldolases share a common ancestor
physiological function
additional information
physiological function
enzyme catalyzes the Mg2+-dependent 1,1-proton transfer that interconverts the enantiomers of mandelate
physiological function
mandelate racemase catalyzes the interconversion of the enantiomers of mandelate
additional information
development and evaluation of a virtual mutant screening method based on the binding energy in the transition state, the method is beneficial in enzyme rational redesign and helps to better understand the catalytic properties of the enzyme, and it is effective in predicting the trend of mutational effects on catalysis, molecular dynamic simulation, overview
additional information
significance of Ser139 in mandelate racemization, the active site residue S139 forms a hydrogen bond with one carboxyl oxygen of the substrate mandelate, residues S139 and E317 of mandelate racemase have a synergistic effect on transition state stabilization, overview
additional information
-
significance of Ser139 in mandelate racemization, the active site residue S139 forms a hydrogen bond with one carboxyl oxygen of the substrate mandelate, residues S139 and E317 of mandelate racemase have a synergistic effect on transition state stabilization, overview
additional information
the enzyme's 20s loop likely undergoes a significant conformational change upon binding (R)-mandelate
additional information
-
the enzyme's 20s loop likely undergoes a significant conformational change upon binding (R)-mandelate
additional information
enzyme molecular dynamics simulation and molecular docking
additional information
role for the Broensted acid-base catalysts of mandelate racemase in transition state stabilization. Enzyme-catalyzed abstraction of an alpha-proton from a carbon acid substrate with a high pKa. Residues Lys166 and His297 play dual roles in catalysis: they act as Broensted acid-base catalysts, and they stabilize both the enolate moiety and phenyl ring of the altered substrate in the transition state
additional information
the thermal stability of mandelate racemase is investigated through molecular dynamics simulations in the temperature range of 30-90°C
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
the enzyme is composed of three distinct structural domains: an N-terminal capping domain consisting of a three-stranded beta-sheet with an antiparallel four-alpha-helix bundle, a central domain consisting of a (beta/alpha)7 beta-barrel, and a short C-terminal domain composed of external beta-strands
additional information
-
the enzyme is composed of three distinct structural domains: an N-terminal capping domain consisting of a three-stranded beta-sheet with an antiparallel four-alpha-helix bundle, a central domain consisting of a (beta/alpha)7 beta-barrel, and a short C-terminal domain composed of external beta-strands
additional information
-
striking structural similarity of mandelate racemase with muconate lactonizing enzyme
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified recombinant homooctameric wild-type enzyme complexed with two analogues of the putative aci-carboxylate intermediate, benzohydroxamate and Cupferron, X-ray diffraction structure at 2.2 A resolution
wild-type and C92S/C264S/K166C mutant enzymes in complex with inhibitors benzilate, 3,3,3-trifluoro-2-hydroxy-2-(trifluoromethyl)-propanoate and tratronate, hanging drop vapor diffusion method, mixing of 0.002 ml of each protein and reservoir solution, the protein solution contains for 3,3,3-trifluoro-2-hydroxy-2-(trifluoromethyl)-propanoate-enzyme crystals 3.3 mM MgCl2, 1 mM inhibitor, and HEPES, 50 mM, pH 7.5, and for the reservoir solution 20% w/v, 120 mM glycine, 70 mM KNO3, and 0.1 M Tris-HCl, pH 8.0, for tartronate-enzyme crystals , 6.0 mg/ml protein, 3.3 mM MgCl2, 20 mM sodium tartronate, and 50 mM HEPES, pH 7.5, and 14% w/v PEG 1500 , 100 mM glycine, and 0.1 M triethanolamin, pH 8.5, as reservoir solution, and for mutant enzyme-benzilate crystals, mutant protein in 3.3 mM MgCl2, 20 mM benzilate, and 50 mM HEPES, pH 7.5, reservoir solution containing 15% w/v PEG 1500, 150 mM glycine, 50 mM NaCl, and 0.1 M HEPES, pH 7.5, equilibration against 0.5 ml reservoir solution, 21°C, and 50% humidity, X-ray diffraction structure determination and analysis at 1.68-1.89 A resolution
crystallographic evidence for stereospecific alkylation by (R)-alpha-phenylglycidate
-
identification of the active site and possible catalytic residues at 2.5 A resolution
-
X-ray crystal structure of mandelate racemase solved at 2.5 A resolution, reveals that the sescondary, tertiary and quarternary structures of mandelate racemase and muconate lactonizing enzyme are remarkably similar
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
A25V
D270N
structure of D270N with (S)-atrolactate bound in the active site reveals no geometric alterations when compared to the structure of the wild type enzyme complexed with (S)-atrolactate, with the exception that the side chain of His297 is tilted and displaced about 0.5A away from Asn270 and towards the (S)-atrolactate. The turnover number for both (R)-mandelate and (S)-mandelate are reduced 10000fold
F52W
compared to wild-type enzyme the catalytic preference of the mutant enzyme is reversed and catalytic efficiency is reduced. Mutant enzyme exhibits higher affinity for (R)-mandelate than for (S)-mandelate, and a higher turnover number with (S)-mandelate as the substrate, relative to that with (R)-mandelate
F52W/Y54W
compared to wild-type enzyme the catalytic preference of the mutant enzyme is reversed and catalytic efficiency is reduced. Mutant enzyme exhibits higher affinity for (R)-mandelate than for (S)-mandelate, and a higher turnover number with (S)-mandelate as the substrate, relative to that with (R)-mandelate
H297N
site-directed mutagenesis, analysis of ligand binding, kinetics
K166C
site-directed mutagenesis, analysis of ligand binding, kinetics
K166M
site-directed mutagenesis, analysis of ligand binding, kinetics
K166M/H297N
site-directed mutagenesis, analysis of ligand binding, kinetics
S139A
site-directed mutagenesis, the mutation leads to a significant reduction of catalytic efficiency by about 45fold and 60fold in R to S and S to R directions
T24S
V22A
V22F
V22I
V22I/V29I/Y54F
site-directed mutagenesis, the mutant shows 3.5fold greater relative activity as compared to the wild-type enzyme. The enhanced catalytic efficiency mainly arises from the elevated kcat, kinetic analysis
V22I/V29L
V26A
V26A/V29L
V26F
V26I
site-directed mutagenesis, the mutant shows 2fold higher catalytic efficiency towards R-mandelamide than the wild-type enzyme
V26I/Y54V
site-directed mutagenesis, the mutant shows 5.2fold higher catalytic efficiency towards (3R)-3-chloromandelic acid than the wild-type enzyme
V26L
V29A
V29F
V29L
Y54F
Y54Q
compared to wild-type enzyme the catalytic preference of the mutant enzyme is reversed and catalytic efficiency is reduced. Mutant enzyme exhibits higher affinity for (R)-mandelate than for (S)-mandelate, and a higher turnover number with (S)-mandelate as the substrate, relative to that with (R)-mandelate
E317Q
-
E317Q with 3400fold reduced turnover number for (R)-mandelate and 29000fold reduced turnover number for (S)-mandelate. E317Q mutant enzyme does not catalyze detectable elimination of Br- from either enantiomer of p-(bromomethyl)mandelate. E317Q mutant enzyme is irreversibly inactivated by racemic alpha-phenylglycidate at a rate comparable to that measured for wild type enzyme
H297N
K166E
-
K166R retains low level of racemase activity. K166R mutant catalyzes the elimination of Br- from only the (R)-enantiomer of (R,S)-p-(bromomethyl)mandelate
N197A
additional information
A25V
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
A25V
variation of the hydrophobic loop, decrease in catalytic efficiency
T24S
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
T24S
variation of the hydrophobic loop, decrease in catalytic efficiency
V22A
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V22A
variation of the hydrophobic loop, decrease in catalytic efficiency
V22F
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V22F
variation of the hydrophobic loop, decrease in catalytic efficiency
V22I
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V22I
variation of the hydrophobic loop. For mandelate, catalytic efficiency similar to wild-type. For substrate 2-naphthylglycolate, increase in catalytic efficiency
V22I/V29L
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V22I/V29L
variation of the hydrophobic loop, catalytic efficiency similar to wild-type
V26A
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V26A
variation of the hydrophobic loop, catalytic efficiency similar to wild-type
V26A/V29L
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V26A/V29L
variation of the hydrophobic loop, decrease in catalytic efficiency
V26F
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V26F
variation of the hydrophobic loop, decrease in catalytic efficiency
V26L
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V26L
variation of the hydrophobic loop, decrease in catalytic efficiency
V29A
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V29A
variation of the hydrophobic loop, decrease in catalytic efficiency
V29F
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V29F
variation of the hydrophobic loop, decrease in catalytic efficiency
V29L
Catalytic efficiencies (kcat/Km) for all mutants are reduced between 6- and 40fold with the exception of V22I, V26A, V29L, and V22I/V29L which have near wildtype efficiencies with mandelate
V29L
variation of the hydrophobic loop. For mandelate, catalytic efficiency similar to wild-type. For substrate 2-naphthylglycolate, increase in catalytic efficiency
V29L
site-directed mutagenesis, the mutant shows 2fold higher catalytic efficiency towards R-2-naphthylglycolate than the wild-type enzyme
H297N
-
H297N has no detectable mandelate racemase activity. However, H297N catalyzes the stereospecific elimination of Br- from racemic p-(bromomethyl)mandelate to give p-(methyl)benzoylformate in 45% yield at a rate equal to that measured for wild type enzyme
H297N
-
H297N, which is inactive as a racemase catalyzes the stereospecific exchange of the alpha-proton of S- but not R-mandelate with solvent D2O at a rate that is 30% of that of the wild type enzyme
N197A
-
Kcat for (R)-mandelate reduced 30fold, Kcat for (S)-mandelate reduced 179fold relative to wild-type
N197A
-
slight activating effect of sucrose on mutant enzyme efficiency. In presewnce of polymeric viscosogens poly(ethylene glycol) and Ficoll, no effect on turnover number or the ratio of turnover number and Km-value for the wild-type enzyme is observed
N197A
-
turnover number is reduced 30fold for (R)-mandelate and 179fold for (S)-mandelate relative to wild-type enzyme. The ratio of turnover number to Km-value is reduced 208fold for (R)-mandelate and 556fold for (S)-mandelate, 3.5fold reduction in affinity for the substrate analogue (R)-atrolactate, but 51fold and 18fold reduction in affinity for alpha-hydroxybenzylphosphonate and benzohydroxamate, respectively
additional information
development and evaluation of a virtual mutant screening method for non-natural substrates based on the binding energy in the transition state, the method is beneficial in enzyme rational redesign and helps to better understand the catalytic properties of the enzyme, and it is effective in predicting the trend of mutational effects on catalysis, molecular dynamic simulation, overview
additional information
Comparison of the binding affinities of the mutant variants with the intermediate/transition state analogues benzohydroxamate and cyclohexanecarbohydroxamate reveals that cationPi/N-Pi interactions between His297 and the hydroxamate/hydroximate moiety and the phenyl ring of benzohydroxamate contribute approximately 0.26 and 0.91 kcal/mol to binding, respectively, while interactions with Lys166 contribute approximately 1.74 and 1.74 kcal/mol, respectively. Lys166 contributes over 2.93 kcal/mol to the binding of (R)-atrolactate, and His297 contributes 2.46 kcal/mol to the binding of (S)-atrolactate
additional information
directed evolution of mandelate racemase by a high-throughput screening method, overview
additional information
evaluation of design of mandelate racemase with higher stability, usage of structural enzyme analysis for reengineering of mandelate racemase for enhanced thermal stability
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
the thermal stability of mandelate racemase is investigated through molecular dynamics simulations in the temperature range of 30-90°C, structural alterations at increasing temperatures, overview. Radius of gyration, surface accessibility, and secondary structure content suggest the instability of mandelate racemase at high temperatures
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
recombinant enzyme exhibits only a slight loss of activity when the enzyme is dialyzed without 200 mM NaCl at 4°C for 8 h, a loss of 20% of its activity when the enzyme is stored for 1 h on ice in the absence of BSA (but stable in the presence of 0.1% BSA), and a 30% reduction in Vmax after storage for 20 days at -70°C in the presence of 200 mM NaCl
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purification of recombinant MR as a fusion protein with an N-terminal hexahistidine tag using immobilized-nickel ion affinity chromatography and elution with a linear gradient of EDTA
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
recombinant Strep-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by affinity chromatography and dialysis
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene mdlA, genetic library construction, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
overexpression of Strep-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
Recombinant MR from is overproduced from Escherichia coli strain BL21(DE3) cells transformed with a pET-15b plasmid containing the MR open reading frame. This construct encodes the MR gene product with an N-terminal hexahistidine tag. Mutants are produced by site-directed mutagenesis
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
analysis
-
direct kinetic assay for mandelate racemase using circular dichroic measurement
biotechnology
-
the demonstrated application of a membrane bioreactor will be a useful method for large-scale dynamic kinetic resulution (DKR) of mandelic acid and for possible other bioconversions in organic media
synthesis
-
DEAE-cellulose-immobilized mandelate racemase can be efficiently used in repeated batch reactions for the racemization of (R)-mandelic acid under mild conditions
additional information
mandelate racemase from Pseudomonas putida is a promising candidate for the dynamic kinetic resolution of alpha-hydroxy carboxylic acids
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Neidhart, D.J.; Kenyon, G.L.; Gerlt, J.A.; Petsko, G.A.
Mandelate racemase and muconate lactonizing enzyme are mechanistically distinct and structurally homologous
Nature
347
692-694
1990
Pseudomonas putida
Tsou, A.Y.; Ransom, S.C.; Gerlt, J.A.; Buechter, D.D.; Babbitt, P.C.; Kenyon, G.L.
Mandelate pathway of Pseudomonas putida: sequence relationships involving mandelate racemase, (S)-mandelate dehydrogenase, and benzoylformate decarboxylase and expression of benzoylformate decarboxylase in Escherichia coli
Biochemistry
29
9856-9862
1990
Pseudomonas putida
Neidhart, D.J.; Powers, V.M.; Kenyon, G.L.; Tsou, A.Y.; Ransom, S.C.; Gerlt, J.A.; Petsko, G.A.
Preliminary X-ray data on crystals of mandelate racemase
J. Biol. Chem.
263
9268-9270
1988
Pseudomonas putida
Ransom, S.C.; Gerlt, J.A.; Powers, V.M.; Kenyon, G.L.
Cloning, DNA sequence analysis, and expression in Escherichia coli of the gene for mandelate racemase from Pseudomonas putida
Biochemistry
27
540-545
1988
Pseudomonas putida
Lin, D.T.; Powers, V.M.; Reynolds, L.J.; Whitman, C.P.; Kozarich, J.W.; Kenyon, G.L.
Evidence for the generation of alpha-carboxy-alpha-hydroxy-p-xylylene from p-(bromomethyl)mandelate by mandelate racemase
J. Am. Chem. Soc.
110
323-324
1988
Pseudomonas putida
-
Whitman, C.P.; Hegeman, G.D.; Cleland, W.W.; Kenyon, G.L.
Symmetry and asymmetry in mandelate racemase catalysis
Biochemistry
24
3936-3942
1985
Pseudomonas putida
Kenyon, G.L.; Hegeman, G.D.
Mandelate racemase
Adv. Enzymol. Relat. Areas Mol. Biol.
50
325-360
1979
Pseudomonas putida
Kenyon, G.L.; Hegeman, G.D.
Mandelate racemase
Methods Enzymol.
46
541-548
1977
Pseudomonas putida
Maggio, E.T.; Kenyon, G.L.; Mildvan, A.S.; Hegeman, G.D.
Mandelate racemase from Pseudomonas putida. Magnetic resonance and kinetic studies of the mechanism of catalysis
Biochemistry
14
1131-1139
1975
Pseudomonas putida
Fee, J.A.; Hegeman, G.D.; Kenyon, G.L.
Mandelate racemase from Pseudomonas putida. Affinity labeling of the enzyme by D,L-alpha-phenylglycidate in the presence of magnesium ion
Biochemistry
13
2533-2538
1974
Pseudomonas putida
Fee, J.A.; Hegeman, G.D.; Kenyon, G.L.
Mandelate racemase from pseudomonas putida. Subunit composition and absolute divalent metal ion requirement
Biochemistry
13
2528-2532
1974
Pseudomonas putida
Hegeman, G.D.
Mandelate racemase (Pseudomonas putida)
Methods Enzymol.
17
670-674
1970
Pseudomonas putida
-
Hegeman, G.D.; Rosenberg, E.Y.; Kenyon, G.L.
Mandelic acid racemase from Pseudomonas putida. Purification and properties of the enzyme
Biochemistry
9
4029-4036
1970
Pseudomonas putida, Pseudomonas putida A.3.12
Schafer, S.L.; Barrett, W.C.; Kallarakal, A.T.; Mitra, B.; Kozarich, J.W.; Gerlt, J.A.
Mechanism of the reaction catalyzed by mandelate racemase: structure and mechanistic properties of the D270N mutant
Biochemistry
35
5662-5669
1996
Pseudomonas putida (P11444), Pseudomonas putida
Sharp, T.R.; Hegeman, G.D.; Kenyon, G.L.
A direct kinetic assay for mandelate racemase using circular dichroic measurements
Anal. Biochem.
94
329-334
1979
Pseudomonas putida
Sharp, T.R.; Hegeman, G.D.; Kenyon, G.L.
Mandelate racemase from Pseudomonas putida. Absence of detectable intermolecular proton transfer accompanying racemization
Biochemistry
16
1123-1128
1977
Pseudomonas putida, Pseudomonas putida A.3.12
Kenyon, G.L.; Hegeman, G.D.
Mandelic acid racemase from Pseudomonas putida. Evidence favoring a carbanion intermediate in the mechanism of action
Biochemistry
9
4036-4043
1970
Pseudomonas putida
Tsou, A.Y.; Ransom, S.C.; Gerlt, J.A.; Powers, V.M.; Kenyon, G.L.
Selection and characterization of a mutant of the cloned gene for mandelate racemase that confers resistance to an affinity label by greatly enhanced production of enzyme
Biochemistry
28
969-975
1989
Pseudomonas putida
Neidhart, D.C.; Howell, P.L.; Petsko, G.A.; Gerlt, J.A.; Kozarich, J.W.; Powers, V.M.; Kenyon, G.L.
Restructuring catalysis in the mandelate pathway
Biochem. Soc. Symp.
57
135-141
1990
Pseudomonas putida
Landro, J.A.; Kallarakal, A.T.; Ransom, S.C.; Gerlt, J.A.; Kozarich, J.W.; Neidhart, D.J.; Kenyon, G.L.
Mechanism of the reaction catalyzed by mandelate racemase. 3. Asymmetry in reactions catalyzed by the H297N mutant
Biochemistry
30
9274-9281
1991
Pseudomonas putida
Neidhart, D.J.; Howell, P.L.; Petsko, G.A.; Powers, V.M.; Li, R.; Kenyon, G.L.; Gerlt, J.A.
Mechanism of the reaction catalyzed by mandelate racemase. 2. Crystal structure of mandelate racemase at 2.5-A resolution: identification of the active site and possible catalytic residues
Biochemistry
30
9264-9273
1991
Pseudomonas putida
Powers, V.M.; Koo, C.W.; Kenyon, G.L.; Gerlt, J.A.; Kozarich, J.W.
Mechanism of the reaction catalyzed by mandelate racemase. 1. Chemical and kinetic evidence for a two-base mechanism
Biochemistry
30
9255-9263
1991
Pseudomonas putida
Gerlt, J.A.; Kenyon, G.L.; Kozarich, J.W.; Neidhart, D.J.; Petsko, G.A.; Powers, V.M.
Mandelate racemase and class-related enzymes
Curr. Opin. Struct. Biol.
2
736-742
1992
Pseudomonas putida
-
Landro, J.A.; Gerlt, J.A.; Kozarich, J.W.; Koo, C.W.; Shah, V.J.; Kenyon, G.L.; Neidhart, D.J.; Fujita, S.; Petsko, G.A.
The role of lysine 166 in the mechanism of mandelate racemase from Pseudomonas putida: mechanistic and crystallographic evidence for stereospecific alkylation by (R)-alpha-phenylglycidate
Biochemistry
33
635-643
1994
Pseudomonas putida
Li, R.; Powers, V.M.; Kozarich, J.W.; Kenyon, G.L.
Racemization of vinylglycolate catalyzed by mandelate racemase
J. Org. Chem.
60
3347-3351
1995
Pseudomonas putida
-
Kallarakal, A.T.; Mitra, B.; Kozarich, J.W.; Gerlt, J.A.; Clifton, J.G.; Petsko, G.A.; Kenyon, G.L.
Mechanism of the reaction catalyzed by mandelate racemase: structure and mechanistic properties of the K166R mutant
Biochemistry
34
2788-2797
1995
Pseudomonas putida
Stecher, H.; Felfer, U.; Faber, K.
Large-scale production of mandelate racemase by Pseudomonas putida ATCC 12633: optimization of enzyme induction and development of a stable crude enzyme preparation
J. Biotechnol.
56
33-40
1997
Pseudomonas putida
-
Mitra, B.; Kallarakal, A.T.; Kozarich, J.W.; Gerlt, J.A.; Clifton, J.G.; Petsko, G.A.; Kenyon, G.L.
Mechanism of the reaction catalyzed by mandelate racemase: importance of electrophilic catalysis by glutamic acid 317
Biochemistry
34
2777-2787
1995
Pseudomonas putida
ST.Maurice, M.; Bearne, S.L.
Reaction intermediate analogues for mandelate racemase: interaction between Asn 197 and the alpha-hydroxyl of the substrate promotes catalysis
Biochemistry
39
13324-13335
2000
Pseudomonas putida
St Maurice, M.; Bearne, S.L.
Kinetics and thermodynamics of mandelate racemase catalysis
Biochemistry
41
4048-4058
2002
Pseudomonas putida
St Maurice, M.; Bearne, S.L.
Hydrophobic nature of the active site of mandelate racemase
Biochemistry
43
2524-2532
2004
Pseudomonas putida (P11444), Pseudomonas putida
Strauss, U.T.; Kandelbauer, A.; Faber, K.
Stabilization and activity-enhancement of mandelate racemase from Pseudomonas putida ATCC 12336 by immobilization
Biotechnol. Lett.
22
515-520
2000
Pseudomonas putida, Pseudomonas putida ATCC 12336
-
Garcia-Viloca, M.; Gonzalez-Lafont, A.; Lluch, J.M.
A QM/MM study of the racemization of vinylglycolate catalyzed by mandelate racemase enzyme
J. Am. Chem. Soc.
123
709-721
2001
Pseudomonas putida (P11444)
Goriup, M.; Strauss, U.T.; Felfer, U.; Kroutil, W.; Faber, K.
Substrate spectrum of mandelate racemase. Part 1: Variation of the a-hydroxy acid moiety
J. Mol. Catal. B
15
207-211
2001
Pseudomonas putida
-
Felfer, U.; Strauss, U.T.; Kroutil, W.; Fabian, W.M.F.; Faber, K.
Substrate spectrum of mandelate racemase. Part 2. (Hetero)-aryl-substituted mandelate derivatives and modulation of activity
J. Mol. Catal. B
15
213-222
2001
Pseudomonas putida
-
Bauer, C.; Boy, M.; Faber, K.; Felfer, U.; Voss, H.
Activation of mandelate racemase via immobilization in lyotropic liquid crystals for biocatalysis in organic solvents: application and modeling
J. Mol. Catal. B
16
91-100
2001
Pseudomonas putida
-
Felfer, U.; Goriup, M.; Koegl, M.F.; Wagner, U.; Larissegger-Schnell, B.; Faber, K.; Kroutil, W.
The substrate spectrum of mandelate racemase: Minimum structural requirements for substrates and substrate model
Adv. Synth. Catal.
347
951-961
2005
Pseudomonas putida
-
Siddiqi, F.; Bourque, J.R.; Jiang, H.; Gardner, M.; St Maurice, M.; Blouin, C.; Bearne, S.L.
Perturbing the hydrophobic pocket of mandelate racemase to probe phenyl motion during catalysis
Biochemistry
44
9013-9021
2005
Pseudomonas putida (P11444), Pseudomonas putida
Burley, R.K.; Bearne, S.L.
Inhibition of mandelate racemase by the substrate-intermediate-product analogue 1,1-diphenyl-1-hydroxymethylphosphonate
Bioorg. Med. Chem. Lett.
15
4342-4344
2005
Pseudomonas putida
Brosseau, C.L.; St.Maurice, M.; Bearne, S.L.; Roscoe, S.G.
Electrochemical quartz crystal nanobalance (EQCN) studies of the adsorption behaviour of an enzyme, mandelate racemase, and its substrate, mandelic acid, on Pt
Electrochim. Acta
50
1289-1297
2005
Pseudomonas putida
-
Prat-Resina, X.; Gonzalez-Lafont, A.; Lluch, J.M.
Reaction mechanism of the mandelate anion racemization catalyzed by mandelate racemase enzyme: A QM/MM Molecular Dynamics Free Energy Study
J. Phys. Chem. B
109
21089-21101
2005
Pseudomonas putida (P11444)
Bourque, J.R.; Burley, R.K.; Bearne, S.L.
Intermediate analogue inhibitors of mandelate racemase: N-Hydroxyformanilide and cupferron
Bioorg. Med. Chem. Lett.
17
105-108
2007
Pseudomonas putida
Choi, W.J.; Lee, K.Y.; Kang, S.H.; Lee, S.B.
Biocatalytic enantioconvergent separation of racemic mandelic acid
Sep. Purif. Technol.
53
178-182
2007
Pseudomonas putida
-
Bourque, J.R.; Bearne, S.L.
Mutational analysis of the active site flap (20s loop) of mandelate racemase
Biochemistry
47
566-578
2008
Pseudomonas putida (P11444), Pseudomonas putida
Narmandakh, A.; Bearne, S.L.
Purification of recombinant mandelate racemase: improved catalytic activity
Protein Expr. Purif.
69
39-46
2010
Pseudomonas putida (P11444), Pseudomonas putida
Nagar, M.; Narmandakh, A.; Khalak, Y.; Bearne, S.L.
Redefining the minimal substrate tolerance of mandelate racemase. Racemization of trifluorolactate
Biochemistry
50
8846-8852
2011
Pseudomonas putida (P11444), Pseudomonas putida
Lietzan, A.D.; Nagar, M.; Pellmann, E.A.; Bourque, J.R.; Bearne, S.L.; St Maurice, M.
Structure of mandelate racemase with bound intermediate analogues benzohydroxamate and cupferron
Biochemistry
51
1160-1170
2012
Pseudomonas putida (P11444), Pseudomonas putida
Nagar, M.; Lietzan, A.D.; St Maurice, M.; Bearne, S.L.
Potent inhibition of mandelate racemase by a fluorinated substrate-product analogue with a novel binding mode
Biochemistry
53
1169-1178
2014
Pseudomonas putida (P11444), Pseudomonas putida
Gu, J.; Liu, M.; Guo, F.; Xie, W.; Lu, W.; Ye, L.; Chen, Z.; Yuan, S.; Yu, H.
Virtual screening of mandelate racemase mutants with enhanced activity based on binding energy in the transition state
Enzyme Microb. Technol.
55
121-127
2014
Pseudomonas putida (P11444)
Gu, J.; Yu, H.
The role of residue S139 of mandelate racemase: synergistic effect of S139 and E317 on transition state stabilization
J. Biomol. Struct. Dyn.
30
585-593
2012
Pseudomonas putida (P11444), Pseudomonas putida
Meshach Paul, D.; Chadah, T.; Senthilkumar, B.; Sethumadhavan, R.; Rajasekaran, R.
Structural distortions due to missense mutations in human formylglycine-generating enzyme leading to multiple sulfatase deficiency
J. Biomol. Struct. Dyn.
36
3575-3585
2017
Pseudomonas putida (P11444)
Yang, C.; Ye, L.; Gu, J.; Yang, X.; Li, A.; Yu, H.
Directed evolution of mandelate racemase by a novel high-throughput screening method
Appl. Microbiol. Biotechnol.
101
1063-1072
2017
Pseudomonas putida (P11444), Pseudomonas putida ATCC 12633 (P11444)
Nagar, M.; Wyatt, B.N.; St Maurice, M.; Bearne, S.L.
Inactivation of mandelate racemase by 3-hydroxypyruvate reveals a potential mechanistic link between enzyme superfamilies
Biochemistry
54
2747-2757
2015
Pseudomonas putida (P11444)
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