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Information on EC 5.3.3.1 - steroid DELTA-isomerase and Organism(s) Comamonas testosteroni and UniProt Accession P00947
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5.3.3.1 steroid DELTA-isomeraseIUBMB Comments
This activity is catalysed by several distinct enzymes (cf. EC 1.1.3.6, cholesterol oxidase and EC 1.1.1.145, 3-hydroxy-5-steroid dehydrogenase).
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Word Map
- 5.3.3.1
-
steroidogenic
- progesterone
- adrenal
-
steroidogenesis
- androgen
- p450scc
- leydig
- star
- testicular
- pregnenolone
- 17beta-hsd
- lh
- aromatase
-
5-3-ketosteroids
- gonad
- testosteroni
-
granulosa
- dehydroepiandrosterone
- luteal
- androstenedione
-
17beta-hydroxysteroid
- dheas
-
theca
- p450arom
- 17alpha-hydroxylase
-
neurosteroids
- cyp11a1
- trilostane
- 19-nortestosterone
-
dienolate
-
5alpha-reductase
- equilenin
-
low-barrier
- 17alpha-hydroxylase/17,20-lyase
-
interna
- 3betahsd
- delta5-3beta-hydroxysteroid
-
intracrine
-
double-bond
- hsd3b1
-
talalay
- 20alpha-hsd
-
beta-proton
The taxonomic range for the selected organisms is: Comamonas testosteroni
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Synonyms
3beta-hsd, ketosteroid isomerase, delta 5-3-ketosteroid isomerase, steroid isomerase, 3beta-hydroxysteroid dehydrogenase/isomerase, steroid delta-isomerase, gst a3-3, 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase, 3beta-hsd/isomerase, 5-ene-4-ene isomerase, 
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topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
3-Keto-DELTA5-steroid isomerase
-
-
-
-
3-Ketosteroid DELTA5-->DELTA4-isomerase
-
-
-
-
3-Oxo steroid DELTA4-DELTA5-isomerase
-
-
-
-
3-Oxo-delta5 steroid isomerase
-
-
-
-
3-Oxosteroid DELTA4-DELTA5-isomerase
-
-
-
-
3-Oxosteroid DELTA5-DELTA4-isomerase
-
-
-
-
3-Oxosteroid isomerase
-
-
-
-
5-Ene-4-ene isomerase
-
-
-
-
5-Pregnene-3,20-dione isomerase
-
-
-
-
delta 5-3-ketosteroid isomerase
-
-
-
-
DELTA-3-ketosteroid isomerase
-
-
-
-
Delta-5-3-ketosteroid isomerase
-
-
-
-
DELTA5(or DELTA4)-3-keto steroid isomerase
-
-
-
-
DELTA5-3-keto steroid isomerase
DELTA5-3-ketosteroid isomerase
-
-
-
-
DELTA5-3-oxosteroid isomerase
-
-
-
-
DELTA5-ketosteroid isomerase
-
-
-
-
DELTA5-steroid isomerase
-
-
-
-
Hydroxysteroid isomerase
-
-
-
-
Isomerase, steroid DELTA
-
-
-
-
Steroid 5-->4-isomerase
-
-
-
-
Steroid isomerase
-
-
-
-
DELTA5-3-keto steroid isomerase
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
a 3-oxo-DELTA5-steroid = a 3-oxo-DELTA4-steroid
mechanism in which the transition state for enolization is dienolate-like, characterized by relatively little proton transfer from Tyr14 in the transition state, and the intermediate in the overall reaction is dienol-like. An alternative mechanism in which the intermediate is stabilized by a short, strong hydrogen bond can also be consistent with the data
-
a 3-oxo-DELTA5-steroid = a 3-oxo-DELTA4-steroid
"flip-flop" mechanism may be involved
-
a 3-oxo-DELTA5-steroid = a 3-oxo-DELTA4-steroid
catalytic mechanism, phenolate binding to the oxyanion hole of ketosteroid isomerase via hydrogen bonding, electrostatic contributions and geometric and electrostatic changes, overview, the KSI reaction involves changes in both geometry and charge distribution as the reaction proceeds from its ground state to its intermediate, Tyr16 and Asp103 are important in catalysis
-
a 3-oxo-DELTA5-steroid = a 3-oxo-DELTA4-steroid
enzyme enhances the coupled motion/hydrogen tunneling contribution to the rate acceleration over the solution reaction
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
isomerization
-
-
-
-
intramolecular oxidoreduction
-
-
-
-
PATHWAY SOURCE
PATHWAYS
KEGG
-
-, -, -, -, -, -, -, -, -, -
SYSTEMATIC NAME
IUBMB Comments
3-oxosteroid DELTA5-DELTA4-isomerase
This activity is catalysed by several distinct enzymes (cf. EC 1.1.3.6, cholesterol oxidase and EC 1.1.1.145, 3-hydroxy-5-steroid dehydrogenase).
CAS REGISTRY NUMBER
COMMENTARY
9031-36-1
-
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
5(10)-estrene-3,17-dione
estr-4-en-3,17-dione
nonsticky substrate
-
?
5-Pregnene-3,20-dione
4-Pregnene-3,20-dione
-
-
-
-
?
androst-5-ene-3,17-dione
androst-4-ene-3,17-dione
-
alpha-secondary deuterium kinetic isotope effects at C6 of the steroid. Presence of coupled motion/hydrogen tunneling in the enzyme-catalyzed reaction
-
-
r
5-Androstene-3,17-dione
4-Androstene-3,17-dione
-
-
-
?
17beta-Hydroxy-5(10)-estren-3-one
?
-
at 0.25% of the activity compared to 5-androstene-3,17-dione
-
-
?
5-Androstene-3,17-dione
4-Androstene-3,17-dione
-
-
-
-
?
5-Androstene-3,17-dione
4-Androstene-3,17-dione
-
residue Asp38 acts as a base to abstract a proton from C-4 of the substrate to form an intermediate dienolate, which is then reprotonated on C-6
-
?
a 3-oxo-DELTA5-steroid
a 3-oxo-DELTA4-steroid
-
phenolates binding to the oxyanion hole of the enzyme via electrostatic interactions, different binding of transition state analogue and substrate, hydrogen bonds shorten with increasing charge localization, overview
-
-
r
additional information
?
-
the 2.3 A structure indicates that Phe101 is not a catalytic residue and that Tyr14 and Asp99 COOH should be directly involved in the stabilization of the dienolate intermediate
-
?
additional information
?
-
-
the 2.3 A structure indicates that Phe101 is not a catalytic residue and that Tyr14 and Asp99 COOH should be directly involved in the stabilization of the dienolate intermediate
-
?
additional information
?
-
the reaction occurs by a two-step general acid-base mechanism involving a dienolate intermediate. Hybrid quantum/classical molecular dynamics simulations elucidating the geometrical, conformational, and electrostatic changes occurring during the isomerization reaction catalyzed by KSI, calculation of rate constants and modeling of the KSI active site, overview. Electrostaic and conformational changes during the proton transfer reactions, rearrangements, overview
-
-
?
additional information
?
-
-
properties of the steroid-enzyme aduct
-
-
?
additional information
?
-
-
equilenin geometrically and electrostatically resembles the dienolate reaction intermediate and transition state, binding structure to enzyme mutant D40N, overview
-
-
?
additional information
?
-
-
mechanism of KSI-catalyzed steroid isomerization, overview
-
-
?
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
5-Androstene-3,17-dione
4-Androstene-3,17-dione
-
-
-
?
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
(17S)-Spiro[5alpha-androstane-17,2'-oxiran]-3-one
-
potent irreversible active-site directed
(17S)-Spiro[estra-1,3,5(10),6,8-pentaene-17,2'-oxiran]-3-ol
-
active-site-directed, irreversible
(17S)-Spiro[estra-1,3,5(10)-triene-17,2'-oxiran]-3-ol
-
potent irreversible active-site directed
10beta-(1-Oxoprop-2-ynyl)oestr-4-ene-3,17-dione
-
irreversible, time-dependent, active-site directed
17beta-(1-Oxoprop-2-ynyl)androst-4-en-3-one
-
irreversible, time-dependent, active-site directed
17beta-Hydroxy-5,10-seco-oestr-4-yne-3,10-dione
-
irreversible, time-dependent, active-site directed
5,10-Seco-oestr-4-yne-3,10,17-trione
-
irreversible, time-dependent, active-site directed
equilenin
-
reaction intermediate analogue, binds to the active site, binding structure, overview
Halothane
-
allosteric modulation of dynamics-function relationship without direct competition, halothane occupancy at the dimer interface disrupts intersubunit hydrogen bonding
Spiro-17beta-oxiranyl-DELTA4-androsten-3-one(4beta)
-
active site-directed, irreversible
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Spiro-17beta-oxiranylestra-1,3,5(10),6,8-pentaene-3-ol
-
active site-directed, irreversible
additional information
-
overview: inactivation by active-site directed acetylenic steroids
-
additional information
-
overview: interaction of the enzyme with steroids
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0486
5-androstene-3,17-dione
25°C, pH 7.0, mutant enzyme F116W
0.067
5-androstene-3,17-dione
-
25°C, pH 6.5, solvent D2O, mutant enzyme D38A
0.073
5-androstene-3,17-dione
-
25°C, pH 8.5, solvent D2O, mutant enzyme D38A
0.085
5-androstene-3,17-dione
-
25°C, pH 7.5, solvent D2O, mutant enzyme D38A
0.094
5-androstene-3,17-dione
-
25°C, pH 8.5, solvent H2O, mutant enzyme D38A
0.096
5-androstene-3,17-dione
-
25°C, pH 7.5, solvent H2O, mutant enzyme D38A
0.099
5-androstene-3,17-dione
-
25°C, pH 6.5, solvent H2O, mutant enzyme D38A
additional information
additional information
thermodynamic parameters and rate constants for isomerization of 5-androstene-3,17-dione
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
230
5-androstene-3,17-dione
-
25°C, pH 6.5, solvent D2O, mutant enzyme D38A
260
5-androstene-3,17-dione
-
25°C, pH 8.5, solvent D2O, mutant enzyme D38A
279
5-androstene-3,17-dione
-
25°C, pH 6.5, solvent H2O, mutant enzyme D38A
340
5-androstene-3,17-dione
-
25°C, pH 6.5, solvent H2O, mutant enzyme D38A
412
5-androstene-3,17-dione
-
25°C, pH 7.5 or 8.5, solvent H2O, mutant enzyme D38A
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.5 - 9
-
pH 7.5: about 40% of maximal activity, pH 9.0: about 60% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
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
11000
-
2 * 11000, SDS-PAGE. At concentrations below 1 mg/ml the enzyme exists as a dimer of MW 26800, determined by gel filtration and ultracentrifugation studies. Concentration-dependent association occurs at higher enzyme concentrations
26000 - 68000
-
protein concentration 0.043 mg/ml-15.6 mg/ml. At concentrations below 1 mg/ml the enzyme exists as a dimer of MW 26800, determined by gel filtration and ultracentrifugation studies. Concentration-dependent association occurs at higher enzyme concentrations. The latter value represents the weight avarage molecular weight of two or more polymerization species in rapid equilibrium, rather than a discrete polymeric form of the enzyme
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
dimer
-
2 * 11000, SDS-PAGE. At concentrations below 1 mg/ml the enzyme exists as a dimer of MW 26800, determined by gel filtration and ultracentrifugation studies. Concentration-dependent association occurs at higher enzyme concentrations
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystals are grown at 12°C from1.3 M ammonium sulfate, 3-5% poly(ethylene glycol)400 and 0.1 M HEPES, pH 7.5 in hanging drops. The crystal structure at 2.3 resolution reveals that the active site environment of the Comamonas testosteroni enzyme is nearly identical to that of Pseudomonas putida enzyme
crystals of mutant enzyme F116W are grown by hanging-drop method
The 2.26 A crystal structure of the enzyme in complex with a reaction intermediate analogue equilenin reveals that both the Tyr14 OH and the Asp99 COOH provide direct hydrogen bonds to the oxyanion of equilenin
1.2-1.5 A resolution X-ray crystallography, 1H and 19F NMR spectroscopy, quantum mechanical calculations, and transition-state analogue binding measurements of the active site. Packing and binding interactions within the KSI active site can constrain local side-chain reorientation and prevent hydrogen bond shortening by 0.1 A or less. This constraint has substantial energetic effects on ligand binding and stabilization of negative charge within the oxyanion hole. Structural features of the oxyanion hole suggest that hydrogen bond formation to the reacting substrate is geometrically optimal in the transition state but not in the ground state. During steroid isomerization, the hybridization of the substrate oxygen changes from a planar sp2 carbonyl to a tetrahedral sp3 dienolate, altering the spatial distribution of its lone pair electrons. This reorientation of atomic orbitals about the substrate oxygen alters its geometric preference for accepting hydrogen bonds
-
study on backbone dynamics in free enzyme and its complex with a steroid analogue, 19-nortestosterone hemisuccinate. Mutation Y14F induces a substantial decrease in the order parameters in free enzyme, indicating that the backbone structures become significantly mobile by mutation, while the chemical shift analysis indicates that the structural perturbations are more profound than those of wild-type upon 19-nortestosterone hemisuccinate binding. In the 19-nortestosterone hemisuccinate complexed mutant, the key active site residues including Tyr14, Asp38 and Asp99 or the regions around them remain flexible with significantly reduced S2 values, whereas the S2 values for many of the residues in the mutant enzyme become even greater than those of wild-type
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D38E
compared to wild-type, 200fold reduction in kcat-value. Analysis of thermodynamic parameters
D99L
the mutant shows about 10000fold decreased kcat compared to the wild type enzyme
F116W
the turnover-number for 5-androstene-3,17-dione is lowered 4.42fold, the KM-value is 3.1fold lower than the Km-value of the wild-type enzyme
Y14F
the mutant shows 100000fold decreased kcat compared to the wild type enzyme
Y14F/D99L
the mutant shows 1000000fold decreased kcat compared to the wild type enzyme
D103A
-
site-directed mutagenesis of the catalytic residue, the mutant shows 5000fold reduced activity compared tot he wild-type enzyme
D38A
-
the catalytic turnover number is 140fold less than that for the wild-type
D38E
-
the mutant gives similar free energies to the native enzyme, with catalytic constants approximately 200-300times less than in wild type enzyme
D40N
-
site-directed mutagenesis, the mutation mimics the protonated aspartate found in the intermediate and equilenin complexes and leads to tighter binding of phenolate and other intermediate analogs
D99A
-
secondary kinetic isotope effects similar to wild-type. Mutation does not significantly decrease the contribution of coupled motion/hydrogen tunneling to the enzymatic reaction
Y14F
Y16F
-
site-directed mutagenesis of the catalytic residue, the mutant shows 50000fold reduced activity compared tot he wild-type enzyme
Y55F/Y88F
-
replacement of Tyr14 by 3-fluorotyrosine in the Y55,88F modified form of the isomerase results in a 4-fold decrease in turnover number
Y57F/D40N
-
site-directed mutagenesis, 3,4,5-trifluorophenol bind as ionized phenolate to KSI containing the Y57F mutation
Y14F
-
secondary kinetic isotope effects similar to wild-type. Mutation does not significantly decrease the contribution of coupled motion/hydrogen tunneling to the enzymatic reaction
Y14F
-
study on backbone dynamics in free enzyme and its complex with a steroid analogue, 19-nortestosterone hemisuccinate. Mutation induces a substantial decrease in the order parameters in free enzyme, indicating that the backbone structures become significantly mobile by mutation, while the chemical shift analysis indicates that the structural perturbations are more profound than those of wild-type upon 19-nortestosterone hemisuccinate binding. In the 19-nortestosterone hemisuccinate complexed mutant, the key active site residues including Tyr14, Asp38 and Asp99 or the regions around them remain flexible with significantly reduced S2 values, whereas the S2 values for many of the residues in the mutant enzyme become even greater than those of wild-type
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
active-site-directed photoinactivation is sensitized by unsaturated steroid ketone photoaffinity reagents
-
in the equilibrium unfolding process the enzyme stability increases by 2.5 kcal/mol in presence of 5% 2,2,2-trifluoroethanol, the increase in entropy by 2,2,2-trifluoroethanol is partially responsible for the increase in stability
-
photoinactivation by ultraviolet irradiation in the presence of the solid-phase photoaffinity reagent DELTA6-testosterone agarose
-
ultraviolet light-dependent photoinactivation is stimulated by DELTA4-3-ketosteroids. The loss of enzymic activity can be correlated with destruction of a single Asp or Asn
-
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
urea
-
the recovery of the activity by refolding is not diminished even after a prolonged time, 24 h of exposure to 7 M urea. Reversibility of the folding is assessed by a 100fold dilution of the denatured protein and subsequent determination of enzyme activity. The refolding kinetics as monitored by fluorescence intensity can be described as a fast first-order process followed by a second-order and a subsequent slow first-order processes, there may be a monomeric folding intermediate
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant C-terminally His6-tagged enzyme from Escherichia coli strain BLR(DE3) to homogeneity by expanded bed adsorption column utilizing immobilized metal affinity chromatography with denaturant concentration in the feed stream from 8 to 0 M urea, chemical extraction from cells, method optimization and validation, overview
recombinant C-terminally His6-tagged enzyme from Escherichia coli strain BLR(DE3) to homogeneity by expanded bed adsorption column utilizing immobilized metal affinity chromatography, this integrated process greatly simplifies the recovery and purification of inclusion body proteins by removing the need for mechanical cell disruption, repeated inclusion body centrifugation, and difficult clarification operations, chemical extraction is as effective as homogenization at releasing the inclusion body proteins from the bacterial cells, over 99% purity of the enzyme, method optimization and validation, overview
recombinant enzyme from Escherichia coli strain BLR(DE3) to homogeneity by nickel affinity chromatography after dilution refolding, method optimization, overview
recombinant wild-type and mutant enzymes from Escherichia coli strain DH5alpha to over 99% purity
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression of C-terminally His6-tagged enzyme as insoluble protein in inclusion bodies in Escherichia coli strain BLR(DE3)
D38A, D38A/Y14F, D38A/Y55F and D38A/D99N proteins are expressed in Epicurian Coli XL1-Blue cells
-
expression of wild-type and mutant enzymes in Escherichia coli strain DH5alpha
-
RENATURED/Commentary
ORGANISM
UNIPROT
LITERATURE
70-87% refolding of recombinant C-terminally His6-tagged enzyme after expression in Escherichia coli as insoluble protein and solubilization by denaturation, detergent treatment, and 40fold dilution by a single-step column-based refolding step with an elution gradient, dilution with buffer 1% bovine serum albumin, 20 mM phosphate, 150 mM NaCl, optimal at pH 6.8, method optimization, overview
dilution refolding of recombinant enzyme from inclusion bodies, denatured recombinant C-terminally His6-tagged enzyme 10fold with refolding buffer containing 50 mM potassium phosphate, pH 7.0, 4°C, incubation at 4°C for 12 h, urea 0.8 M
repeated adsorptive refolding in an expanded bed with gradient change in the feed-stream composition from denaturing buffer containing 8 M urea and 50 mM potassium phosphate, pH 6.8 to the refolding buffer containing 50 mM potassium phosphate, pH 6.8, constant liquid feed rate, method evaluation, overview
the recovery of the activity by refolding is not diminished even after a prolonged time, 24 h of exposure to 7 M urea
-
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Kawahara, F.S.; Wang, S.F.; Talalay, P.
The preparation and properties of crystalline DELTA5-3-ketosteroid isomerase
J. Biol. Chem.
237
1500-1506
1962
Comamonas testosteroni
Jones, J.B.; Ship, S.
Inhibition of the 3-ketosteroid DELTA5->DELTA4 isomerase of Pseudomonas testosteroni by some bromo-3-ketosteroid derivatives
Biochim. Biophys. Acta
258
800-809
1972
Comamonas testosteroni
Geynet, Ph.; Gallay, J.; Alfsen, A.
Mammalian 3-oxosteroid DELTA5-DELTA4-isomerase. Solubilization by calcium ions and kinetic characteristics toward C19 and C21 steroids
Eur. J. Biochem.
31
464-469
1972
Comamonas testosteroni
Tivol, W.F.; Beckman, E.D.; Benisek, W.F.
Effect of protein concentration on the molecular weight of DELTA5-3-ketosteroid isomerase
J. Biol. Chem.
250
271-275
1975
Comamonas testosteroni
Benson, A.M.; Suruda, A.J.; Talalay, P.
Concentration-dependent association of DELTA5-3-ketosteroid isomerase of Pseudomonas testosteroni
J. Biol. Chem.
250
276-280
1975
Comamonas testosteroni
Benson, A.M.; Suruda, A.J.; Barrack, E.R.; Talalay, P.
Steroid-transforming enzymes
Methods Enzymol.
34B
557-566
1974
Comamonas testosteroni
Westbrook, E.M.
Characterization of a hexagonal crystal form of an enzyme of steroid metabolism, DELTA5-3-ketosteroid isomerase: a new method of crystal density measurement
J. Mol. Biol.
103
659-664
1976
Comamonas testosteroni
Weintraub, H.; Vincent, F.; Baulieu, E.E.; Alfsen, A.
Interaction of steroids with Pseudomonas testosteroni 3-oxosteroid DELTA4-DELTA5 isomerase
Biochemistry
16
5045-5053
1977
Comamonas testosteroni
Penning, T.M.; Covey, D.F.; Talalay, P.
Inactivation of DELTA5-3-oxo steroid isomerase with active-site-directed acetylenic steroids
Biochem. J.
193
217-227
1981
Comamonas testosteroni
Penning, T.M.; Covey, D.F.; Talalay, P.
Irreversible inactivation of DELTA5-3-ketosteroid isomerase of Pseudomonas testosteroni by acetylenic suicide substrates. Mechanism of formation and properties of the steroid-enzyme adduct
J. Biol. Chem.
256
6842-6850
1981
Comamonas testosteroni
Hearne, M.; Benisek, W.F.
Use of a solid-phase photoaffinity reagent to label a steroid binding site: application to the DELTA5-3-ketosteroid isomerase of Pseudomonas testosteroni
Biochemistry
24
7511-7516
1985
Comamonas testosteroni
Westbrook, E.M.; Sigler, P.B.; Berman, H.; Glusker, J.P.; Bunick, G.; Benson, A.; Talalay, P.
Characterization of a monoclinic crystal form of an enzyme of steroid metabolism, DELTA5-3-ketosteroid isomerase
J. Mol. Biol.
103
665-667
1976
Comamonas testosteroni
Brooks, B.; Benisek, W.F.
Mechanism of the reaction catalyzed by DELTA5-3-ketosteroid isomerase of Comamonas (Pseudomonas) testosteroni: kinetic properties of a modified enzyme in which tyrosine 14 is replaced by 3-fluorotyrosine
Biochemistry
33
2682-2687
1994
Comamonas testosteroni
Pollack, R.M.; Bantia, S.; Bounds, P.L.; Koffman, B.M.
pH-Dependence of the kinetic parameters for 3-oxo-DELTA5-steroid isomerase substrate catalysis and inhibition by (3S)-spiro[5alpha-androstane-3,2'-oxiran]-17-one
Biochemistry
25
1905-1911
1986
Comamonas testosteroni
Bevins, C.L.; Pollack, R.M.; Kayser, R.H.; Bounds, P.L.
Detection of transient enzyme-steroid complex during active-site-directed irreversible inhibition of 3-oxo-DELTA5-steroid isomerase
Biochemistry
25
5159-5164
1986
Comamonas testosteroni
Cho, H.S.; Choi, G.; Choi, K.Y.; Oh, B.H.
Crystal structure and enzyme mechanism of Delta 5-3-ketosteroid isomerase from Pseudomonas testosteroni
Biochemistry
37
8325-8330
1998
Comamonas testosteroni (P00947), Comamonas testosteroni
Benisek, W.F.
Labeling of DELTA-3-ketosteroid isomerase by photoexcited steroid ketones
Methods Enzymol.
46
469-479
1977
Comamonas testosteroni
Kayser, R.H.; Bounds, P.L.; Bevins, C.L.; Pollack, R.M.
Affinity alkylation of bacterial DELTA5-3-ketosteroid isomerase. Identification of the amino acid modified by steroidal 17beta-oxiranes
J. Biol. Chem.
258
909-915
1983
Comamonas testosteroni
Covey, D.F.; Robinson, C.H.
Conjugated allenic 3-oxo-5,10-secosteroids. Irreversible inhibitors of DELTA5-3-ketosteroid isomerase
J. Am. Chem. Soc.
98
5038-5040
1976
Comamonas testosteroni
Bazold, F.H.; Robinson, C.H.
Irreversible inhibition of DELTA5-3-ketosteroid isomerase by 5,10-secosteroids
J. Am. Chem. Soc.
97
2575-2578
1975
Comamonas testosteroni
-
Hearne, M.; Benisek, W.F.
Modification of DELTA5-3-ketosteroid isomerase induced by ultraviolet irradiation in the presence of the solid-phase photoaffinity reagent DELTA6-testosterone agarose
J. Protein Chem.
3
87-97
1984
Comamonas testosteroni
-
Kim, D.H.; Jang, D.S.; Nam, G.H.; Yun, S.; Cho, J.H.; Choi, G.; Lee, H.C.; Choi, K.Y.
Equilibrium and kinetic analysis of folding of ketosteroid isomerase from Comamonas testosteroni
Biochemistry
39
13084-13092
2000
Comamonas testosteroni
Henot, F.; Pollack, R.M.
Catalytic activity of the D38A mutant of 3-oxo-DELTA 5-steroid isomerase: recruitment of aspartate-99 as the base
Biochemistry
39
3351-3359
2000
Comamonas testosteroni
Cho, H.S.; Ha, N.C.; Choi, G.; Kim, H.J.; Lee, D.; Oh, K.S.; Kim, K.S.; Lee, W.; Choi, K.Y.; Oh, B.H.
Crystal structure of DELTA(5)-3-ketosteroid isomerase from Pseudomonas testosteroni in complex with equilenin settles the correct hydrogen bonding scheme for transition state stabilization
J. Biol. Chem.
274
32863-32868
1999
Comamonas testosteroni (P00947)
Yun, S.; Jang do, S.; Choi, G.; Kim, K.S.; Choi, K.Y.; Lee, H.C.
Trifluoroethanol increases the stability of DELTA(5)-3-ketosteroid isomerase. 15N NMR relaxation studies
J. Biol. Chem.
277
23414-23419
2002
Comamonas testosteroni
Yun, Y.S.; Lee, T.H.; Nam, G.H.; Jang, D.S.; Shin, S.; Oh, B.H.; Choi, K.Y.
Origin of the different pH activity profile in two homologous ketosteroid isomerases
J. Biol. Chem.
278
28229-28236
2003
Comamonas testosteroni (P00947), Comamonas testosteroni
Yonkunas, M.J.; Xu, Y.; Tang, P.
Anesthetic interaction with ketosteroid isomerase: Insights from molecular dynamics simulations
Biophys. J.
89
2350-2356
2005
Comamonas testosteroni
Houck, W.J.; Pollack, R.M.
Temperature effects on the catalytic activity of the D38E mutant of 3-oxo-DELTA5-steroid isomerase: favorable enthalpies and entropies of activation relative to the nonenzymatic reaction catalyzed by acetate ion
J. Am. Chem. Soc.
126
16416-16425
2004
Comamonas testosteroni (P00947)
Hutchinson, M.H.; Chase, H.A.
Refolding strategies for ketosteroid isomerase following insoluble expression in Escherichia coli
Biotechnol. Bioeng.
94
1089-1098
2006
Comamonas testosteroni (P00947)
Hutchinson, M.H.; Morreale, G.; Middelberg, A.P.; Chase, H.A.
Production of enzymatically active ketosteroid isomerase following insoluble expression in Escherichia coli
Biotechnol. Bioeng.
95
724-733
2006
Comamonas testosteroni (P00947)
Hutchinson, M.H.; Chase, H.A.
Intensified process for the purification of an enzyme from inclusion bodies using integrated expanded bed adsorption and refolding
Biotechnol. Prog.
22
1187-1193
2006
Comamonas testosteroni (P00947)
Kraut, D.A.; Sigala, P.A.; Pybus, B.; Liu, C.W.; Ringe, D.; Petsko, G.A.; Herschlag, D.
Testing electrostatic complementarity in enzyme catalysis: hydrogen bonding in the ketosteroid isomerase oxyanion hole
PLoS Biol.
4
501-519
2006
Comamonas testosteroni, Pseudomonas putida
-
Sigala, P.A.; Kraut, D.A.; Caaveiro, J.M.; Pybus, B.; Ruben, E.A.; Ringe, D.; Petsko, G.A.; Herschlag, D.
Testing geometrical discrimination within an enzyme active site: constrained hydrogen bonding in the ketosteroid isomerase oxyanion hole
J. Am. Chem. Soc.
130
13696-13708
2008
Comamonas testosteroni, Pseudomonas putida (P07445)
Wilde, T.C.; Blotny, G.; Pollack, R.M.
Experimental evidence for enzyme-enhanced coupled motion/quantum mechanical hydrogen tunneling by ketosteroid isomerase
J. Am. Chem. Soc.
130
6577-6585
2008
Comamonas testosteroni
Lee, H.J.; Yoon, Y.J.; Jang, D.S.; Kim, C.; Cha, H.J.; Hong, B.H.; Choi, K.Y.; Lee, H.C.
15N NMR relaxation studies of Y14F mutant of ketosteroid isomerase: the influence of mutation on backbone mobility
J. Biochem.
144
159-166
2008
Comamonas testosteroni
Chakravorty, D.; Soudackov, A.; Hammes-Schiffer, S.
Hybrid quantum/classical molecular dynamics simulations of the proton transfer reactions catalyzed by ketosteroid isomerase: Analysis of hydrogen bonding, conformational motions, and electrostatics
Biochemistry
48
10608-10619
2009
Comamonas testosteroni (P00947)
Sigala, P.; Caaveiro, J.; Ringe, D.; Petsko, G.; Herschlag, D.
Hydrogen bond coupling in the ketosteroid isomerase active site
Biochemistry
48
6932-6939
2009
Comamonas testosteroni, Pseudomonas putida (P07445)
Chakravorty, D.K.; Hammes-Schiffer, S.
Impact of mutation on proton transfer reactions in ketosteroid isomerase: insights from molecular dynamics simulations
J. Am. Chem. Soc.
132
7549-7555
2010
Comamonas testosteroni (P00947)
van der Kamp, M.W.; Chaudret, R.; Mulholland, A.J.
QM/MM modelling of ketosteroid isomerase reactivity indicates that active site closure is integral to catalysis
FEBS J.
280
3120-3131
2013
Comamonas testosteroni
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