Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary extracted from

  • Truong, L.; Hevener, K.E.; Rice, A.J.; Patel, K.; Johnson, M.E.; Lee, H.
    High-level expression, purification, and characterization of Staphylococcus aureus dihydroorotase (PyrC) as a cleavable His-SUMO fusion (2013), Protein Expr. Purif., 88, 98-106.
    View publication on PubMedView publication on EuropePMC

Application

EC Number Application Comment Organism
3.5.2.3 drug development the essential enzyme is a target for antibacterial drug design Staphylococcus aureus

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.5.2.3 gene pyrC, DNA and amino acid sequence determination and analysis, sequence comparison, expression of N-terminal His-SUMO-tagged enzyme in Escherichia coli strain BL21(DE3) with an HRV 3C protease recognition site inserted between the SUMO tag and SaPyrC to allow for improved cleavage by HRV protease, method optimization Staphylococcus aureus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.5.2.3 additional information
-
 additional information forward and reverse kinetic rate constants for both tagged and wild-type enzyme, kinetic analysis, overview Staphylococcus aureus
3.5.2.3 0.118
-
 (S)-dihydroorotate pH 8.3, 25°C, recombinant detagged enzyme Staphylococcus aureus
3.5.2.3 0.167
-
 (S)-dihydroorotate pH 8.3, 25°C, recombinant tagged enzyme Staphylococcus aureus
3.5.2.3 0.334
-
 N-carbamoyl-L-aspartate pH 8.3, 25°C, recombinant tagged enzyme Staphylococcus aureus
3.5.2.3 0.348
-
 N-carbamoyl-L-aspartate pH 8.3, 25°C, recombinant detagged enzyme Staphylococcus aureus

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.5.2.3 Zn2+ dihydroorotase enzyme is a di-metalloenzyme that is dependent upon two zinc atoms in the active site for activity Staphylococcus aureus

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
3.5.2.3 46370
-
 2 * 46370, sequence calculation Staphylococcus aureus
3.5.2.3 46500
-
 detagged recombinant enzyme, gel filtration Staphylococcus aureus
3.5.2.3 60700
-
 His-SUMO-tagged recombinant enzyme, gel filtration Staphylococcus aureus

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
3.5.2.3 (S)-dihydroorotate + H2O Staphylococcus aureus
-
 N-carbamoyl-L-aspartate
-
 r
3.5.2.3 (S)-dihydroorotate + H2O Staphylococcus aureus MW2
-
 N-carbamoyl-L-aspartate
-
 r

Organism

EC Number Organism UniProt Comment Textmining
3.5.2.3 Staphylococcus aureus P65907 methicillin-resistant strain, gene pyrC
-
3.5.2.3 Staphylococcus aureus MW2 P65907 methicillin-resistant strain, gene pyrC
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.5.2.3 recombinant N-terminal His-SUMO-tagged enzyme SaPyrC from Escherichia coli strain BL21(DE3), by nickel affinity chromatography, dialysis, and His-SUMO tag cleavage through HRV 3C protease, another step of nickel affinity chromatography to remove the tag, followed by ultrafiltration, and gel filtration, to about 95% purity, method optimization, the His-SUMO tag affects enzyme activity slightly Staphylococcus aureus

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.5.2.3 0.68
-
 purified recombinant detagged enzyme, pH 8.3, 25°C Staphylococcus aureus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.2.3 (S)-dihydroorotate + H2O
-
 Staphylococcus aureus N-carbamoyl-L-aspartate
-
 r
3.5.2.3 (S)-dihydroorotate + H2O
-
 Staphylococcus aureus MW2 N-carbamoyl-L-aspartate
-
 r

Subunits

EC Number Subunits Comment Organism
3.5.2.3 homodimer 2 * 46370, sequence calculation Staphylococcus aureus
3.5.2.3 More the His-SUMO tag does not interfere with SaPyrC dimerization Staphylococcus aureus

Synonyms

EC Number Synonyms Comment Organism
3.5.2.3 pyrC
-
 Staphylococcus aureus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.5.2.3 25
-
 assay at Staphylococcus aureus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.5.2.3 1.55
-
 (S)-dihydroorotate pH 8.3, 25°C, recombinant detagged enzyme Staphylococcus aureus
3.5.2.3 1.87
-
 (S)-dihydroorotate pH 8.3, 25°C, recombinant tagged enzyme Staphylococcus aureus
3.5.2.3 1.9
-
 N-carbamoyl-L-aspartate pH 8.3, 25°C, recombinant tagged enzyme Staphylococcus aureus
3.5.2.3 2.48
-
 N-carbamoyl-L-aspartate pH 8.3, 25°C, recombinant detagged enzyme Staphylococcus aureus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.5.2.3 6
-
 cyclization reaction Staphylococcus aureus
3.5.2.3 9
-
 hydrolysis reaction Staphylococcus aureus

General Information

EC Number General Information Comment Organism
3.5.2.3 metabolism third enzyme in the bacterial de novo pyrimidine biosynthesis pathway, overview Staphylococcus aureus

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.5.2.3 5.69
-
 N-carbamoyl-L-aspartate pH 8.3, 25°C, recombinant tagged enzyme Staphylococcus aureus
3.5.2.3 7.13
-
 N-carbamoyl-L-aspartate pH 8.3, 25°C, recombinant detagged enzyme Staphylococcus aureus
3.5.2.3 11.2
-
 (S)-dihydroorotate pH 8.3, 25°C, recombinant tagged enzyme Staphylococcus aureus
3.5.2.3 13.1
-
 (S)-dihydroorotate pH 8.3, 25°C, recombinant detagged enzyme Staphylococcus aureus