Literature summary for 2.5.1.18 extracted from

Wang, B.; Peng, Y.; Zhang, T.; Ding, J.
Crystal structures and kinetic studies of human Kappa class glutathione transferase provide insights into the catalytic mechanism (2011), Biochem. J., 439, 215-225.

Cloned(Commentary)

Cloned (Comment)	Organism
expression of His-tagged enzyme in Escherichia coli	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant hGSTk in apo-form and in complex with S-hexylglutathione, hanging drop vapour diffusion method, apo-form of hGSTk in 20 mM NaH2PO4, pH 7.4, 20 mM NaCl, 1 mM EDTA and 7.2 mM 2-mercaptoethanol, hGSTk in complex in 20 mM HEPES, pH 7.0, 50 mM NaCl, 1 mM EDTA and 1 mM DTT, supplemented with S-hexylglutathione at a molar ratio of 1:2, is mixed with an equal volume of reservoir solution containing 0.2 M NaSCN and 20% PEG3350, 20°C, X-ray diffraction structure determination and analysis at 1.8-1.9 A resolution	Homo sapiens

Crystallization (Comment)

Organism

purified recombinant hGSTk in apo-form and in complex with S-hexylglutathione, hanging drop vapour diffusion method, apo-form of hGSTk in 20 mM NaH2PO4, pH 7.4, 20 mM NaCl, 1 mM EDTA and 7.2 mM 2-mercaptoethanol, hGSTk in complex in 20 mM HEPES, pH 7.0, 50 mM NaCl, 1 mM EDTA and 1 mM DTT, supplemented with S-hexylglutathione at a molar ratio of 1:2, is mixed with an equal volume of reservoir solution containing 0.2 M NaSCN and 20% PEG3350, 20°C, X-ray diffraction structure determination and analysis at 1.8-1.9 A resolution

Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
D201A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
D69A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
D69T	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
F83A	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
F83W	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
F87A	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
F87W	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
I44A	site-directed mutagenesis, H-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
K62A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
K62E	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
L15A	site-directed mutagenesis, H-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
L183A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
L58A	site-directed mutagenesis, H-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
L88A	site-directed mutagenesis, H-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
L92A	site-directed mutagenesis, H-site mutation, inactive mutant	Homo sapiens
M66A	site-directed mutagenesis, H-site mutation, inactive mutant	Homo sapiens
M91A	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
M91K	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
N53A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
P17A	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
P55A	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
P56A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
P56A	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
R202A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
S16A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
S19A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
S200A	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
W126A	site-directed mutagenesis, H-site mutation, the mutant shows increased activity compared to the wild-type enzyme	Homo sapiens
Y18A	site-directed mutagenesis, G-site mutation, inactive mutant	Homo sapiens
Y18F	site-directed mutagenesis, G-site mutation, the mutant shows reduced activity compared to the wild-type enzyme	Homo sapiens
Y18H	site-directed mutagenesis, G-site mutation, inactive mutant	Homo sapiens
Y18L	site-directed mutagenesis, G-site mutation, inactive mutant	Homo sapiens

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
additional information	-	additional information	recombinant hGSTk, steady-state kinetics, overview	Homo sapiens
0.17	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant W126A	Homo sapiens
0.19	-	glutathione	pH 7.4, 25°C, recombinant wild-type GSTk	Homo sapiens
0.34	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P17A	Homo sapiens
0.62	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant M91A	Homo sapiens
0.63	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant M91K	Homo sapiens
0.64	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F83W	Homo sapiens
0.74	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P56A	Homo sapiens
0.91	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F87A	Homo sapiens
1.03	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S200A	Homo sapiens
1.08	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P55A	Homo sapiens
1.1	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant wild-type GSTk	Homo sapiens
1.25	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant K62A	Homo sapiens
1.3	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F83A	Homo sapiens
1.38	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F87W	Homo sapiens
1.45	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L58A	Homo sapiens
1.61	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S19A	Homo sapiens
1.68	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant P56A	Homo sapiens
2.01	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant I44A	Homo sapiens
2.59	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L15A	Homo sapiens
4.44	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L88A	Homo sapiens
4.53	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant N53A	Homo sapiens
4.74	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D201A	Homo sapiens
9.81	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant L183A	Homo sapiens
17.34	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S16A	Homo sapiens
20.45	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant Y18F	Homo sapiens
25.26	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant K62E	Homo sapiens
60.89	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D69T	Homo sapiens
105.4	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D69A	Homo sapiens
123.6	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant R202A	Homo sapiens

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
mitochondrial matrix	-	Homo sapiens	5759	-
peroxisome	matrix	Homo sapiens	5777	-

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	-	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His-tagged enzyme from Escherichia coli by nickel affinity chromatography and dialysis	Homo sapiens

Reaction

Reaction	Comment	Organism	Reaction ID
RX + glutathione = HX + R-S-glutathione	catalytic mechanism of kappa class GST, overview. Substrate binding induces a conformational change of the active site from an open conformation in the apo-form to a closed conformation in the S-hexylglutathione-bound complex, facilitating formations of the G site (GSH-binding site) and the H site (hydrophobic substrate-binding site). The conserved Ser16 at the G site functions as the catalytic residue in the deprotonation of the thiol group and the conserved Asp69, Ser200,Asp201 andArg202 form a network of interactions with gamma -glutamyl carboxylate to stabilize the thiolate anion. The H site is a large hydrophobic pocket with conformational flexibility to allow the binding of different hydrophobic substrates. The kinetic mechanism of hGSTk conforms to a rapid equilibrium random sequential Bi Bi model	Homo sapiens	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
1-chloro-2,4-dinitrobenzene + glutathione	-	Homo sapiens	2,4-dinitrophenyl-glutathione + HCl	-	?

Subunits

Subunits	Comment	Organism
dimer	crystal structure	Homo sapiens

Synonyms

Synonyms	Comment	Organism
GSTk	-	Homo sapiens
kappa class glutathione transferase	-	Homo sapiens
Kappa class GST	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
25	-	assay at	Homo sapiens

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism
0.17	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S16A	Homo sapiens
0.32	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D69A	Homo sapiens
0.47	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D201A	Homo sapiens
0.47	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant L183A	Homo sapiens
0.67	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant R202A	Homo sapiens
0.75	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant K62E	Homo sapiens
0.89	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant K62A	Homo sapiens
0.99	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S200A	Homo sapiens
1.18	-	glutathione	pH 7.4, 25°C, recombinant wild-type GSTk	Homo sapiens
1.19	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L15A	Homo sapiens
1.33	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D69T	Homo sapiens
1.56	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant Y18F	Homo sapiens
1.75	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant N53A	Homo sapiens
2.29	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant P56A	Homo sapiens
2.38	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P17A	Homo sapiens
2.4	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L58A	Homo sapiens
2.94	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant W126A	Homo sapiens
3.18	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F83W	Homo sapiens
3.28	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant I44A	Homo sapiens
3.74	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S19A	Homo sapiens
3.94	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant wild-type GSTk	Homo sapiens
4.18	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F87A	Homo sapiens
4.5	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P55A	Homo sapiens
5	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L88A	Homo sapiens
5.11	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant M91K	Homo sapiens
5.5	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant M91A	Homo sapiens
5.78	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F87W	Homo sapiens
6.24	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F83A	Homo sapiens
7.83	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P56A	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.4	-	assay at	Homo sapiens

General Information

General Information	Comment	Organism
evolution	kappa class GSTs are a distinct class of the GST superfamily or Trx (thioredoxin)-fold superfamily because of their unique cellular localization, function and structure. GSTks do not contain a conserved CXXC motif at the active site	Homo sapiens
additional information	G-site and H-site structure analysis and modeling, overview	Homo sapiens
physiological function	glutathione transferases are a family of enzymes that primarily catalyse nucleophilic addition of the thiol of GSH to a variety of hydrophobic electrophiles in the cellular detoxification of cytotoxic and genotoxic compounds	Homo sapiens

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism
0.00304	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D69A	Homo sapiens
0.00542	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant R202A	Homo sapiens
0.0098	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S16A	Homo sapiens
0.0218	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D69T	Homo sapiens
0.0297	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant K62E	Homo sapiens
0.0479	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant L183A	Homo sapiens
0.0763	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant Y18F	Homo sapiens
0.0992	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant D201A	Homo sapiens
0.232	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S19A	Homo sapiens
0.386	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant N53A	Homo sapiens
0.712	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant K62A	Homo sapiens
0.737	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L15A	Homo sapiens
0.961	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant S200A	Homo sapiens
1.13	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L88A	Homo sapiens
1.36	-	glutathione	pH 7.4, 25°C, recombinant GSTk mutant P56A	Homo sapiens
1.63	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant I44A	Homo sapiens
1.66	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant L58A	Homo sapiens
3.58	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant wild-type GSTk	Homo sapiens
4.17	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P55A	Homo sapiens
4.19	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F87W	Homo sapiens
4.59	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F87A	Homo sapiens
4.8	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F83A	Homo sapiens
4.97	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant F83W	Homo sapiens
6.21	-	glutathione	pH 7.4, 25°C, recombinant wild-type GSTk	Homo sapiens
7	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P17A	Homo sapiens
8.11	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant M91K	Homo sapiens
8.87	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant M91A	Homo sapiens
10.6	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant P56A	Homo sapiens
17.3	-	1-chloro-2,4-dinitrobenzene	pH 7.4, 25°C, recombinant GSTk mutant W126A	Homo sapiens