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Information on EC 5.5.1.1 - Muconate cycloisomerase and Organism(s) Pseudomonas putida and UniProt Accession P08310
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5.5.1.1 Muconate cycloisomeraseIUBMB Comments
Requires Mn2+. Also acts (in the reverse reaction) on 3-methyl-cis,cis-muconate and, very slowly, on cis,trans-muconate. Not identical with EC 5.5.1.7 (chloromuconate cycloisomerase) or EC 5.5.1.11 (dichloromuconate cycloisomerase).
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Word Map
- 5.5.1.1
- muconolactone
-
beta-ketoadipate
-
cycloisomerization
- protoanemonin
- 5-chloromuconolactone
- maleylacetate
- 2-chloro-cis,cis-muconate
- cutaneum
- cis-dienelactone
- chlorocatechols
- o-succinylbenzoate
- 4-chlorophenol
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5.3.3.4
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3-chloro
- 4-chlorocatechol
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gerlt
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phenol-degrading
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ortho-cleavage
- trans-dienelactone
The taxonomic range for the selected organisms is: Pseudomonas putida
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
Synonyms
muconate cycloisomerase, muconate lactonizing enzyme, mc ii, cis,cis-muconate cycloisomerase, cis,cis-muconate lactonizing enzyme, rho-2-cmci, mle-1, anti-mle, tcmle, cis,cis-muconate-lactonizing enzyme, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
cis,cis-Muconate cycloisomerase
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cis,cis-Muconate-lactonizing enzyme
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Enzymes, muconate-lactonizing
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Isomerase, muconate cyclo-
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MC
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MLE
MLEI
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Muconate cycloisomerase I
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Muconate lactonizing enzyme
TcMLE
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MLE
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Muconate lactonizing enzyme
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
(+)-muconolactone = cis,cis-muconate
a concerted or stepwise mechanism involving carbonium ion or covalent intermediates are not likely to be operative for the enzyme-catalyzed syn (cis) elimination
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(+)-muconolactone = cis,cis-muconate
Mn2+ and Lys167 provide electrophilic stabilization of the enol/enolate intermediate that is formed during the course of the reaction
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(+)-muconolactone = cis,cis-muconate
reaction may proceed through formation of an intermediate carbanion located alpha to a carboxyl group, subsequent protonation of this putative carbanion, the carbanion is generated by attack of the remote carboxylate group on a double bond
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(+)-muconolactone = cis,cis-muconate
in the reverse reaction the alpha proton of muconolactone is abstracted by a homolog of Lys166(Lys169) to generate an enolic intermediate, this intermediate undergoes vinylopous beta-elimination of the remote carboxylate group to produce cis,cis-muconate
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
1,2-elimination
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4,5-elimination
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PATHWAY SOURCE
PATHWAYS
BRENDA
KEGG
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SYSTEMATIC NAME
IUBMB Comments
2,5-Dihydro-5-oxofuran-2-acetate lyase (decyclizing)
Requires Mn2+. Also acts (in the reverse reaction) on 3-methyl-cis,cis-muconate and, very slowly, on cis,trans-muconate. Not identical with EC 5.5.1.7 (chloromuconate cycloisomerase) or EC 5.5.1.11 (dichloromuconate cycloisomerase).
CAS REGISTRY NUMBER
COMMENTARY
9023-72-7
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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
2-chloro-cis,cis-muconate
5-chloromuconolactone + 2-chloromuconolactone
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-
?
2-chloro-cis,cis-muconate
(+)-2-chloromuconolactone + (+)-5-chloromuconolactone
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mutant enzymes I54V and I54V/Y59W form 5-chloromuconolactone almost exclusively. Mutant enzymes F329I and Y59W form 2-chloromuconolactone as the main product. Mutant enzyme A271S hehaves similar to wild-type enzyme
?
3-chloro-cis,cis-muconate
protoanemonin + ?
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formed by wild-type enzyme and mutant enzymes I54V, A271S, K276N, F329I, L333V, I54V/F329I
?
cis,cis-Muconate
(+)-Muconolactone
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-
?
cis,cis-Muconate
(+)-Muconolactone
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i.e. cis,cis-2,4-hexadiendioate, , r
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?
cis,cis-Muconate
(+)-Muconolactone
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i.e. cis,cis-2,4-hexadiendioate, , r
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?
cis,cis-Muconate
?
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the enzyme plays a key role in the beta-ketoadipate pathway of benzoate degradation
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?
cis,cis-Muconate
?
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enzyme of the beta-ketoadipate pathway
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?
cis,cis-muconate
muconolactone
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the enzyme plays a crucial role in the bacterial degradation of aromatic compounds by converting cis,cis-muconate, the product of catechol ring cleavage to (4S)-muconolactone
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?
3-chloro-cis,cis-muconate
additional information
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wild-type enzyme forms protoanemonin and 2-chloroprotoanemonin, mutant enzyme K169A forms cis-dienelactone and 2-chloro-cis-dienelactone
?
additional information
?
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no activity against 2-carboxy-cis,cis-muconate
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?
additional information
?
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the enzyme is a component of the beta-ketoadipate pathway
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?
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
cis,cis-muconate
muconolactone
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the enzyme plays a crucial role in the bacterial degradation of aromatic compounds by converting cis,cis-muconate, the product of catechol ring cleavage to (4S)-muconolactone
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?
additional information
?
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the enzyme is a component of the beta-ketoadipate pathway
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?
cis,cis-Muconate
?
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the enzyme plays a key role in the beta-ketoadipate pathway of benzoate degradation
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?
cis,cis-Muconate
?
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enzyme of the beta-ketoadipate pathway
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?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mn2+
Mn2+
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divalent metal ion required, enzyme contains a single binding site per subunit, Km: 0.003-0.0045 mM
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.045
2,4-dichloro-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme I54V/F329I
0.057
2-methyl-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme A271S or I54V/F329
0.1
3-fluoro-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme A271S or I54V/F329
0.0064
3-methyl-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme A271S or I54V/F329
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
1320
2,4-dichloro-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme I54V/F329I
900
2-chloro-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme Y59W and A271S
84000
2-methyl-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme A271S or I54V/F329
600000
2-methyl-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme K276N or F329I
18600
3-methyl-cis,cis-muconate
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pH 7.5, 25°C, mutant enzyme A271S or I54V/F329
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0035
2,4-dichloro-cis,cis-muconate
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inhibitor of cis,cis-muconate conversion
0.031
2-fluoro-cis,cis-muconate
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inhibitor of cis,cis-muconate conversion
0.095
3-chloro-cis,cis-muconate
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inhibitor of cis,cis-muconate conversion
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
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
40000
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hexamer
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6 * 41600-44000, sedimentation equilibrium studies in presence of 6 M guanidine-HCL, gel filtration in presence of guanidine-HCl, calculation from amino acid composition, SDS-PAGE
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method, packing of the octameric enzyme in the crystal form is unusual, because the asymmetric unit contains three subunits
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
F329I
the ratio of turnover number to Km-value: 5.7fold decrease with cis,cis-muconate as substrate, 1.5fold increase with 2-chloro-cis,cis-muconate, 3.4fold increased with 3-chloro-cis,cis-muconate, 3.16fold increase with 2,4-dichloro-cis,cis-muconate, 1.7fold decrease with 3-fluoro-cis,cis-muconate, 1.55fold increase with 2-methyl-cis,cis-muconate and 5.8fold increase with 3-methyl-cis,cis-muconate
I54V
ratio of turnover number to Km-value: 11.9fold decrease with cis,cis-muconate as substrate, nearly identical with 2-chloro-cis,cis-muconate, 22fold increase with 3-chloro-cis,cis-muconate, 1.6fold decrease with 2,4-dichloro-cis,cis-muconate, 5.75fold decrease with 3-fluoro-cis,cis-muconate, 6fold decrease with 2-methyl-cis,cis-muconate and 1.3fold increase with 3-methyl-cis,cis-muconate
A271S
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ratio of turnover number to Km-value: 1.5fold decrease with cis,cis-muconate as substrate, 3.3fold decrease with 2-chloro-cis,cis-muconate as substrate, 27fold increase with 3-chloro-cis,cis-muconate as substrate, 1.9fold decrease with 2,4-dichloro-cis,cis-muconate as substrate, 1.5fold decrease with 3-fluoro-cis,cis-muconate as substrate, 1.1fold increase with 2-methyl-cis,cis-muconate as substrate, 1.1fold decrease with 3-methyl-cis,cis-muconate as substrate
F329I
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ratio of turnover number to Km-value: 5.7fold decrease with cis,cis-muconate as substrate, 1.6fold increase with 2-chloro-cis,cis-muconate as substrate, 3.4fold increase with 3-chloro-cis,cis-muconate as substrate, 3.2fold increase with 2,4-dichloro-cis,cis-muconate as substrate, 1.7fold decrease with 3-fluoro-cis,cis-muconate as substrate, 1.5fold increase with 2-methyl-cis,cis-muconate as substrate, 5.8fold decrease with 3-methyl-cis,cis-muconate as substrate
I54V
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ratio of turnover number to Km-value: 11.9fold decrease with cis,cis-muconate as substrate, nearly identical to wild-type value with 2-chloro-cis,cis-muconate and 3-methyl-cis,cis-muconate as substrate, 22fold increase with 3-chloro-cis,cis-muconate as substrate, 1.67fold decrease with 2,4-dichloro-cis,cis-muconate as substrate, 5.8fold decrease with 3-fluoro-cis,cis-muconate as substrate, 8.8fold decrease with 2-methyl-cis,cis-muconate as substrate
I54V/F329I
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ratio of turnover number to Km-value: 81.5fold decrease with cis,cis-muconate as substrate, 2.6fold decrease with 2-chloro-cis,cis-muconate as substrate, 2.7fold increase with 3-chloro-cis,cis-muconate as substrate, 3.8fold increase with 2,4-dichloro-cis,cis-muconate as substrate, 2fold decrease with 3-fluoro-cis,cis-muconate as substrate, 4fold decrease with 2-methyl-cis,cis-muconate as substrate, 2fold decrease with 3-methyl-cis,cis-muconate as substrate
K169A
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70000fold reduction in activity with cis,cis-muconate, 110000fold reduction in activity with 3-fluoro-cis,cis-muconate, 310fold reduction in activity with 3-chloro-cis,cis-muconate, 2.6fold reduction in activity with 2,4-dichloro-cis,cis-muconate. Wild-type enzyme forms protoanemonin and 2-chloroprotoanemonin from 3-chloro-cis,cis-muconate, mutant enzyme K169A forms cis-dienelactone and 2-chloro-cis-dienelactone
K276N
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ratio of turnover number to Km-value: 10.7fold decrease with cis,cis-muconate as substrate, 8.7fold decrease with 2-chloro-cis,cis-muconate as substrate, 3.5fold increase with 3-chloro-cis,cis-muconate as substrate, 8.7fold decrease with 2,4-dichloro-cis,cis-muconate as substrate, 4.1fold decrease with 3-fluoro-cis,cis-muconate as substrate, 16.7fold decrease with 2-methyl-cis,cis-muconate as substrate, 12.1fold decrease with 3-methyl-cis,cis-muconate as substrate
L333V
-
ratio of turnover number to Km-value: 238fold decrease with cis,cis-muconate as substrate, 41.5fold decrease with 2-chloro-cis,cis-muconate as substrate, 3.4fold increase with 3-chloro-cis,cis-muconate as substrate, 5.9fold decrease with 2,4-dichloro-cis,cis-muconate as substrate, 1.4fold decrease with 3-fluoro-cis,cis-muconate as substrate, 34.6fold decrease with 2-methyl-cis,cis-muconate as substrate, 16.5fold decrease with 3-methyl-cis,cis-muconate as substrate
Y59W
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ratio of turnover number to Km-value: 12.9fold decrease with cis,cis-muconate as substrate, nearly identical to wild-type value with 2-chloro-cis,cis-muconate as substrate, 4.4fold decrease with 3-fluoro-cis,cis-muconate, 1.8fold decrease with 2-methyl-cis,cis-muconate as substrate, 10.2fold decrease with 3-methyl-cis,cis-muconate as substrate
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Meagher, R.B.; Ornston, L.N.
Relationships among enzymes of the beta-ketoadipate pathway. I. Properties of cis,cis-muconate-lactonizing enzyme and muconolactone isomerase from Pseudomonas putida
Biochemistry
12
3523-3530
1973
Pseudomonas putida
Ornston, L.N.
Conversion of catechol and protocatechuate to beta-ketoadipate (Pseudomonas putida)
Methods Enzymol.
17A
529-549
1970
Pseudomonas putida
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Goldman, A.; Ollis, D.; Ngai, K.L.; Steitz, T.A.
Crystal structure of muconate lactonizing enzyme at 6.5 A resolution
J. Mol. Biol.
182
353-355
1995
Pseudomonas putida
Ngai, K.L.; Ornston, L.N.; Kallen, R.G.
Enzymes of the beta-ketoadipate pathway in Pseudomonas putida: kinetic and magnetic resonance studies of the cis,cis-muconate cycloisomerase catalyzed reaction
Biochemistry
22
5223-5230
1983
Pseudomonas putida
Avigad, G.; Englard, S.; Olsen, B.R.; Wolfenstein-Todel, C.; Wiggins, R.
Molecular properties of cis,cis-muconate cycloisomerase from Pseudomonas putida
J. Mol. Biol.
89
651-662
1974
Pseudomonas putida
Mars, A.E.; Kasberg, T.; Kaschabek, S.R.; van Agteren, M.H.; Janssen, D.B.; Reineke, W.
Microbial degradation of chloroaromatics: use of the meta-cleavage pathway for mineralization of chlorobenzene
J. Bacteriol.
179
4530-4537
1997
Pseudomonas putida
Babbitt, P.C.; Mrachko, G.T.; Hasson, M.S.; Huisman, G.W.; Kolter, R.; Ringe, D.; Petsko, G.A.; Kenyon, G.L.; Gerlt, J.A.
A functionally diverse enzyme superfamily that abstracts the alpha protons of carboxylic acids
Science
267
1159-1161
1995
Pseudomonas putida
Helin, S.; Kahn, P.C.; Guha, B.L.; Mallows, D.G.; Goldman, A.
The refined X-ray structure of muconate lactonizing enzyme from Pseudomonas putida PRS2000 at 1.85 A resolution
J. Mol. Biol.
254
918-941
1995
Pseudomonas putida, Pseudomonas putida PRS2000
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
Meagher, R.B.; Ngai, K-L.; Ornston, L.N.
Muconate cycloisomerase
Methods Enzymol.
188
126-130
1990
Pseudomonas putida
Ngai, K.L.; Kallen, R.G.
Enzymes of the beta-ketoadipate pathway in Pseudomonas putida: primary secondary kinetic and equilibrium deuterium isotope effects upon the interconversion of (+)-muconolactone to cis,cis-muconate catalyzed by cis,cis-muconate cycloisomerase
Biochemistry
22
5231-5236
1993
Pseudomonas putida
Goldman, A.; Ollis, D.L.; Steitz, T.A.
Crystal structure of muconate lactonizing enzyme at 3 A resolution
J. Mol. Biol.
194
143-153
1987
Pseudomonas putida
Aldrich, T.L.; Frantz, B.; Gill, J.F.; Kilbane, J.J.; Chakrabarty, A.M.
Cloning and complete nucleotide sequence determination of the catB gene encoding cis,cis-muconate lactonizing enzyme
Gene
52
185-195
1987
Pseudomonas putida
Oltmanns, R.H.; Rast, H.G.; Reineke, W.
Degradation of 1,4-dichlorobenzene by enriched and constructed bacteria
Appl. Microbiol. Biotechnol.
28
609-616
1988
Alcaligenes sp., Pseudomonas sp., Pseudomonas putida, Pseudomonas putida WR1323, Alcaligenes sp. R3, Pseudomonas sp. RHO1
-
Vollmer, M.D.; Hoier, H.; Hecht, H.J.; Schell, U.; Groning, J.; Goldman, A.; Schlomann, M.
Substrate specificity of and product formation by muconate cycloisomerases: an analysis of wild-type enzymes and engineered variants
Appl. Environ. Microbiol.
64
3290-3299
1998
Acinetobacter calcoaceticus, Pseudomonas putida, Pseudomonas putida PRS2000
Kaulmann, U.; Kaschabek, S.R.; Schlomann, M.
Mechanism of chloride elimination from 3-chloro- and 2,4-dichloro-cis,cis-muconate: new insight obtained from analysis of muconate cycloisomerase variant CatB-K169A
J. Bacteriol.
183
4551-4561
2001
Pseudomonas putida
Hasson, M.S.; Schlichting, I.; Moulai, J.; Taylor, K.; Barrett, W.; Kenyon, G.L.; Babbitt, P.C.; Gerlt, J.A.; Petsko, G.A.; Ringe, D.
Evolution of an enzyme active site: the structure of a new crystal form of muconate lactonizing enzyme compared with mandelate racemase and enolase
Proc. Natl. Acad. Sci. USA
95
10396-10401
1998
Pseudomonas putida
Schell, U.; Helin, S.; Kajander, T.; Schlomann, M.; Goldman, A.
Structural basis for the activity of two muconate cycloisomerase variants toward substituted muconates
Proteins Struct. Funct. Genet.
34
125-136
1999
Pseudomonas putida (P08310)
Gerlt, J.
Evolution of enzymatic activities in the enolase superfamily: Stereochemically distinct mechanisms in two famillies of cis,cis-muconate lactonizing enzymes
Biochemistry
48
1445-1453
2009
Mycolicibacterium smegmatis, Pseudomonas putida, Pseudomonas fluorescens (Q4K9X1), Pseudomonas fluorescens
EXTERNAL LINKS (specific for EC-Number 5.5.1.1)
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