BRENDA - Enzyme Database
show all sequences of 2.3.1.221

Structure and function of an iterative polyketide synthase thioesterase domain catalyzing Claisen cyclization in aflatoxin biosynthesis

Korman, T.P.; Crawford, J.M.; Labonte, J.W.; Newman, A.G.; Wong, J.; Townsend, C.A.; Tsai, S.C.; Proc. Natl. Acad. Sci. USA 107, 6246-6251 (2010)

Data extracted from this reference:

Cloned(Commentary)
Cloned (Commentary)
Organism
expression of recombinant His-tagged thioesterase/Claisen cyclase in Escherichia coli, expression of the selenomethionine-labeled PksA TE in Escherichia coli methionine auxotroph strain B834(DE3)
Aspergillus sp.
Crystallization (Commentary)
Crystallization (Commentary)
Organism
purified recombinant selenomethionine-labeled of PksA TE, sitting drop vapour diffusion method, 5 mg/ml protein in 20 mM Tris-HCl pH 7.5 containing 5% glycerol and 2 mM DTT is mixed with well solution containing 0.2 M ammonium acetate, 0.1 M sodium citrate pH 5.6, and 30% PEG 4000, 25C, 2 days, X-ray diffraction structure determination and analysis, modeling
Aspergillus sp.
Engineering
Protein Variants
Commentary
Organism
D1964N
site-directed mutagenesis of a catalytic residue
Aspergillus sp.
D2070N
site-directed mutagenesis, the mutation results in only a slightly reduced rate of hydrolysis compared to the apo-mutant
Aspergillus sp.
H2088F
site-directed mutagenesis of a catalytic residue
Aspergillus sp.
S1937A
site-directed mutagenesis of a catalytic residue
Aspergillus sp.
Natural Substrates/ Products (Substrates)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
ID
7 malonyl-CoA + hexanoyl-[acyl-carrier protein]
Aspergillus sp.
-
7 CoA + norsolorinic acid anthrone + [acyl-carrier protein] + 7 CO2 + 2 H2O
-
-
?
Organism
Organism
UniProt
Commentary
Textmining
Aspergillus sp.
Q12052
-
-
Purification (Commentary)
Purification (Commentary)
Organism
recombinant His-tagged thioesterase/Claisen cyclase and selenomethionine-labeled PksA TE from Escherichia coli
Aspergillus sp.
Reaction
Reaction
Commentary
Organism
Reaction ID
7 malonyl-CoA + hexanoyl-[acyl-carrier protein] = 7 CoA + norsolorinic acid anthrone + [acyl-carrier protein] + 7 CO2 + 2 H2O
mechanism of thioesterase/Claisen cyclase-catalyzed chain-termination of fungal aromatic polyketide biosynthesis. The ACP of the ACP-bound substrate is displaced upon thioesterase-catalyzed transesterification. Rotation of the substrate side chain can occur once the ACP leaves the pocket, and the thioesterase can then close. Thioesterase conformational constraints as observed in the closed-form crystal structure guide Claisen-type cyclization to release noranthrone, i.e. norsolorinic acid anthrone, the polyketide precursor of aflatoxin B1. Domain structure and reaction mechanism, detailed overview
Aspergillus sp.
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
Substrate Product ID
7 malonyl-CoA + hexanoyl-[acyl-carrier protein]
-
723638
Aspergillus sp.
7 CoA + norsolorinic acid anthrone + [acyl-carrier protein] + 7 CO2 + 2 H2O
-
-
-
?
Subunits
Subunits
Commentary
Organism
More
domain architecture, the enzyme shows an alpha/beta-hydrolase fold in the catalytic closed form with a distinct hydrophobic substrate-binding chamber involving the PksA thioesterase/Claisen cyclase residues Ser1937, His2088, and Asp1964, which constitute the catalytic triad conserved in the alpha/beta-hydrolase family, detailed overview
Aspergillus sp.
Synonyms
Synonyms
Commentary
Organism
PksA
-
Aspergillus sp.
polyketide synthase A
-
Aspergillus sp.
Temperature Optimum [C]
Temperature Optimum [C]
Temperature Optimum Maximum [C]
Commentary
Organism
25
-
assay at
Aspergillus sp.
pH Optimum
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7.5
-
assay at
Aspergillus sp.
Cloned(Commentary) (protein specific)
Commentary
Organism
expression of recombinant His-tagged thioesterase/Claisen cyclase in Escherichia coli, expression of the selenomethionine-labeled PksA TE in Escherichia coli methionine auxotroph strain B834(DE3)
Aspergillus sp.
Crystallization (Commentary) (protein specific)
Crystallization
Organism
purified recombinant selenomethionine-labeled of PksA TE, sitting drop vapour diffusion method, 5 mg/ml protein in 20 mM Tris-HCl pH 7.5 containing 5% glycerol and 2 mM DTT is mixed with well solution containing 0.2 M ammonium acetate, 0.1 M sodium citrate pH 5.6, and 30% PEG 4000, 25C, 2 days, X-ray diffraction structure determination and analysis, modeling
Aspergillus sp.
Engineering (protein specific)
Protein Variants
Commentary
Organism
D1964N
site-directed mutagenesis of a catalytic residue
Aspergillus sp.
D2070N
site-directed mutagenesis, the mutation results in only a slightly reduced rate of hydrolysis compared to the apo-mutant
Aspergillus sp.
H2088F
site-directed mutagenesis of a catalytic residue
Aspergillus sp.
S1937A
site-directed mutagenesis of a catalytic residue
Aspergillus sp.
Natural Substrates/ Products (Substrates) (protein specific)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
ID
7 malonyl-CoA + hexanoyl-[acyl-carrier protein]
Aspergillus sp.
-
7 CoA + norsolorinic acid anthrone + [acyl-carrier protein] + 7 CO2 + 2 H2O
-
-
?
Purification (Commentary) (protein specific)
Commentary
Organism
recombinant His-tagged thioesterase/Claisen cyclase and selenomethionine-labeled PksA TE from Escherichia coli
Aspergillus sp.
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ID
7 malonyl-CoA + hexanoyl-[acyl-carrier protein]
-
723638
Aspergillus sp.
7 CoA + norsolorinic acid anthrone + [acyl-carrier protein] + 7 CO2 + 2 H2O
-
-
-
?
Subunits (protein specific)
Subunits
Commentary
Organism
More
domain architecture, the enzyme shows an alpha/beta-hydrolase fold in the catalytic closed form with a distinct hydrophobic substrate-binding chamber involving the PksA thioesterase/Claisen cyclase residues Ser1937, His2088, and Asp1964, which constitute the catalytic triad conserved in the alpha/beta-hydrolase family, detailed overview
Aspergillus sp.
Temperature Optimum [C] (protein specific)
Temperature Optimum [C]
Temperature Optimum Maximum [C]
Commentary
Organism
25
-
assay at
Aspergillus sp.
pH Optimum (protein specific)
pH Optimum Minimum
pH Optimum Maximum
Commentary
Organism
7.5
-
assay at
Aspergillus sp.
General Information
General Information
Commentary
Organism
metabolism
polyketide synthase A is a multidomain PKS central to the biosynthesis of aflatoxin B1, a potent environmental carcinogen
Aspergillus sp.
additional information
the synthetic versatility of thioesterase domains in fungal nonreducing, iterative PKSs extends to Claisen cyclase chemistry by catalyzing C-C ring closure reactions as opposed to thioester hydrolysis or O-C/N-C macrocyclization observed in other thioesterase structures. Catalysis of C-C bond formation as a product release mechanism dramatically expands the synthetic potential of PKSs, structural analyses of the thioesterase/CLC domain in polyketide synthase A
Aspergillus sp.
General Information (protein specific)
General Information
Commentary
Organism
metabolism
polyketide synthase A is a multidomain PKS central to the biosynthesis of aflatoxin B1, a potent environmental carcinogen
Aspergillus sp.
additional information
the synthetic versatility of thioesterase domains in fungal nonreducing, iterative PKSs extends to Claisen cyclase chemistry by catalyzing C-C ring closure reactions as opposed to thioester hydrolysis or O-C/N-C macrocyclization observed in other thioesterase structures. Catalysis of C-C bond formation as a product release mechanism dramatically expands the synthetic potential of PKSs, structural analyses of the thioesterase/CLC domain in polyketide synthase A
Aspergillus sp.
Other publictions for EC 2.3.1.221
No.
1st author
Pub Med
title
organims
journal
volume
pages
year
Activating Compound
Application
Cloned(Commentary)
Crystallization (Commentary)
Engineering
General Stability
Inhibitors
KM Value [mM]
Localization
Metals/Ions
Molecular Weight [Da]
Natural Substrates/ Products (Substrates)
Organic Solvent Stability
Organism
Oxidation Stability
Posttranslational Modification
Purification (Commentary)
Reaction
Renatured (Commentary)
Source Tissue
Specific Activity [micromol/min/mg]
Storage Stability
Substrates and Products (Substrate)
Subunits
Synonyms
Temperature Optimum [C]
Temperature Range [C]
Temperature Stability [C]
Turnover Number [1/s]
pH Optimum
pH Range
pH Stability
Cofactor
Ki Value [mM]
pI Value
IC50 Value
Activating Compound (protein specific)
Application (protein specific)
Cloned(Commentary) (protein specific)
Cofactor (protein specific)
Crystallization (Commentary) (protein specific)
Engineering (protein specific)
General Stability (protein specific)
IC50 Value (protein specific)
Inhibitors (protein specific)
Ki Value [mM] (protein specific)
KM Value [mM] (protein specific)
Localization (protein specific)
Metals/Ions (protein specific)
Molecular Weight [Da] (protein specific)
Natural Substrates/ Products (Substrates) (protein specific)
Organic Solvent Stability (protein specific)
Oxidation Stability (protein specific)
Posttranslational Modification (protein specific)
Purification (Commentary) (protein specific)
Renatured (Commentary) (protein specific)
Source Tissue (protein specific)
Specific Activity [micromol/min/mg] (protein specific)
Storage Stability (protein specific)
Substrates and Products (Substrate) (protein specific)
Subunits (protein specific)
Temperature Optimum [C] (protein specific)
Temperature Range [C] (protein specific)
Temperature Stability [C] (protein specific)
Turnover Number [1/s] (protein specific)
pH Optimum (protein specific)
pH Range (protein specific)
pH Stability (protein specific)
pI Value (protein specific)
Expression
General Information
General Information (protein specific)
Expression (protein specific)
KCat/KM [mM/s]
KCat/KM [mM/s] (protein specific)
722459
Vagstad
Interrogation of global active ...
Aspergillus parasiticus
J. Am. Chem. Soc.
134
6865-6877
2012
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723638
Korman
Structure and function of an i ...
Aspergillus sp.
Proc. Natl. Acad. Sci. USA
107
6246-6251
2010
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1
4
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723262
Crawford
Structural basis for biosynthe ...
Aspergillus parasiticus
Nature
461
1139-1143
2009
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1
1
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721052
Crawford
Deconstruction of iterative mu ...
Aspergillus parasiticus
Science
320
243-246
2008
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1
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4
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2
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723194
Chang
The Aspergillus parasiticus po ...
Aspergillus parasiticus, Aspergillus parasiticus SRRC 2043 / RHN1
Mol. Gen. Genet.
248
270-277
1995
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