Literature summary for 1.20.4.4 extracted from

Messens, J.; Hayburn, G.; Desmyter, A.; Laus, G.; Wyns, L.
The essential catalytic redox couple in arsenate reductase from Sataphylococcus aureus (1999), Biochemistry, 38, 16857-16865.

Cloned(Commentary)

Cloned (Comment)	Organism
expressed in Escherichia coli	Staphylococcus aureus
expression in Escherichia coli. Wild-type enzyme and the Cys mutants (C15A, C10A, C82A, C82S, C89A, C10SC15S, C10SC15A) are expressed in Escherichia coli. Wild-type enzyme, mutant enzyme C15A, mutant enzyme C10A, mutant enzyme C82S, mutant enzyme C89A, and mutant enzyme C10SC15A are expressed soluble and with high yields. Mutant enzyme C82A is found in inclusion bodies, and the double mutant C10S/C15S is not expressed	Staphylococcus aureus

Cloned (Comment)

Organism

expressed in Escherichia coli

Staphylococcus aureus

expression in Escherichia coli. Wild-type enzyme and the Cys mutants (C15A, C10A, C82A, C82S, C89A, C10SC15S, C10SC15A) are expressed in Escherichia coli. Wild-type enzyme, mutant enzyme C15A, mutant enzyme C10A, mutant enzyme C82S, mutant enzyme C89A, and mutant enzyme C10SC15A are expressed soluble and with high yields. Mutant enzyme C82A is found in inclusion bodies, and the double mutant C10S/C15S is not expressed

Staphylococcus aureus

Protein Variants

Protein Variants	Comment	Organism
C10A	inactive mutant enzyme	Staphylococcus aureus
C10A	site-directed mutagenesis, mutation of Cys 10, 82, and 89 leads to redox-inactive enzymes	Staphylococcus aureus
C10S/C15A	double mutation, no enzymatic activity	Staphylococcus aureus
C10S/C15S	double mutation, no enzymatic activity	Staphylococcus aureus
C10SC15A	inactive mutant enzyme	Staphylococcus aureus
C15A	site-directed mutagenesis, only ArsC wild type and ArsC C15A show enzymatic activity	Staphylococcus aureus
C15A	as compared to wild-type enzyme the affinity is reduced ba a factor of 2	Staphylococcus aureus
C82A	site-directed mutagenesis, mutation of Cys 10, 82, and 89 leads to redox-inactive enzymes	Staphylococcus aureus
C82S	inactive mutant enzyme	Staphylococcus aureus
C82S	site-directed mutagenesis, mutation of Cys 10, 82, and 89 leads to redox-inactive enzymes	Staphylococcus aureus
C89A	inactive mutant enzyme	Staphylococcus aureus
C89A	site-directed mutagenesis, mutation of Cys 10, 82, and 89 leads to redox-inactive enzymes	Staphylococcus aureus
additional information	essential cysteinyl residues and redox couple in arsenate reductase are identified by a combination of site-specific mutagenesis and endoprotease-digest mass spectroscopy analysis	Staphylococcus aureus

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
0.000066	-	arsenate	wild-type	Staphylococcus aureus
0.000066	-	arsenate	pH 7.5, 37°C, wild-type enzyme	Staphylococcus aureus
0.000134	-	arsenate	C15A mutant	Staphylococcus aureus
0.000134	-	arsenate	pH 7.5, 37°C, mutant enzyme C15A	Staphylococcus aureus

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
14400	-	gel filtration	Staphylococcus aureus
21000	-	gel filtration	Staphylococcus aureus

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
arsenate + thioredoxin	Staphylococcus aureus	the enzyme encoded by Staphylococcus aureus arsenic-resistance plasmid pI258 reduces intracellular arsenate to the more toxic arsenite, which is subsequently extruded from the cell	arsenite + thioredoxin disulfide + H2O	-	?
additional information	Staphylococcus aureus	assays are performed with different arsenate concentrations and arsenate reductase concentrations in the presence of 0.42 microM Escherichia coli thioredoxin, 0.14 microM Escherichia coli thioredoxin reductase and 125 microM NADPH	?	-	?

Organism

Organism	UniProt	Comment	Textmining
Staphylococcus aureus	P0A006	-	-

Purification (Commentary)

Purification (Comment)	Organism
-	Staphylococcus aureus
wild-type ArsC and the Cys mutants, gel filtration more than 97% pure	Staphylococcus aureus

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
arsenate + thioredoxin	-	Staphylococcus aureus	arsenite + thioredoxin disulfide + H2O	-	?
arsenate + thioredoxin	the enzyme encoded by Staphylococcus aureus arsenic-resistance plasmid pI258 reduces intracellular arsenate to the more toxic arsenite, which is subsequently extruded from the cell	Staphylococcus aureus	arsenite + thioredoxin disulfide + H2O	-	?
arsenate + thioredoxin	the enzyme uses an intramolecular thiol pair (Cys82, Cys89) for the reduction of arsenate	Staphylococcus aureus	arsenite + thioredoxin disulfide + H2O	-	?
additional information	assays are performed with different arsenate concentrations and arsenate reductase concentrations in the presence of 0.42 microM Escherichia coli thioredoxin, 0.14 microM Escherichia coli thioredoxin reductase and 125 microM NADPH	Staphylococcus aureus	?	-	?

Subunits

Subunits	Comment	Organism
?	x * 14400, SDS-PAGE	Staphylococcus aureus

Synonyms

Synonyms	Comment	Organism
ArsC	-	Staphylococcus aureus
arsenate reductase	-	Staphylococcus aureus

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Staphylococcus aureus

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism
0.061	-	arsenate	C15A ArsC mutant, at 2 microM AsO43-	Staphylococcus aureus
0.061	-	arsenate	pH 7.5, 37°C, mutant enzyme C15A, at 0.002 mM arsenate	Staphylococcus aureus
0.075	-	arsenate	wild-type ArsC, at 2 microM AsO43-	Staphylococcus aureus
0.075	-	arsenate	pH 7.5, 37°C, wild-type enzyme, at 0.002 mM arsenate	Staphylococcus aureus
0.08	-	arsenate	C15A ArsC mutant, at 10 mM AsO43-	Staphylococcus aureus
0.08	-	arsenate	pH 7.5, 37°C, mutant enzyme C15A, at 10 mM arsenate	Staphylococcus aureus
0.165	-	arsenate	wild-type ArsC, at 10 mM AsO43-	Staphylococcus aureus
0.165	-	arsenate	pH 7.5, 37°C, wild-type enzyme, at 10 mM arsenate	Staphylococcus aureus

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Staphylococcus aureus

Cofactor

Cofactor	Comment	Organism	Structure
additional information	ArsC couples to thioredoxin, thioredoxin reductase, and NADPH to be enzymatically active	Staphylococcus aureus

General Information

General Information	Comment	Organism
physiological function	the enzyme encoded by Staphylococcus aureus arsenic-resistance plasmid pI258 reduces intracellular arsenate to the more toxic arsenite, which is subsequently extruded from the cell	Staphylococcus aureus

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism
441	-	arsenate	pH 7.5, 37°C, mutant enzyme C15A, at 0.002 mM arsenate	Staphylococcus aureus
455	-	arsenate	C15A mutant, 2.6 times lower	Staphylococcus aureus
1136	-	arsenate	wild-type	Staphylococcus aureus
1170	-	arsenate	pH 7.5, 37°C, wild-type enzyme, at 0.002 mM arsenate	Staphylococcus aureus