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Literature summary for 2.5.1.72 extracted from

  • Marinoni, I.; Nonnis, S.; Monteferrante, C.; Heathcote, P.; Haertig, E.; Boettger, L.H.; Trautwein, A.X.; Negri, A.; Albertini, A.M.; Tedeschi, G.
    Characterization of L-aspartate oxidase and quinolinate synthase from Bacillus subtilis (2008), FEBS J., 275, 5090-5107.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expression with C-terminal His-tag Bacillus subtilis

Protein Variants

Protein Variants Comment Organism
C110S 0.4 mol iron per mol of protein, no enzymic activity Bacillus subtilis
C230S 0.6 mol iron per mol of protein, no enzymic activity Bacillus subtilis
C259S 4.5 mol iron per mol of protein, 80% of wild-type activity Bacillus subtilis
C318S 3.3 mol iron per mol of protein, 75% of wild-type activity Bacillus subtilis
C318S/C320S 0.3 mol iron per mol of protein, no enzymic activity Bacillus subtilis
C320S 1.5 mol iron per mol of protein, no enzymic activity Bacillus subtilis
C82S 4.3 mol iron per mol of protein, activity similar to wild-type Bacillus subtilis

Inhibitors

Inhibitors Comment Organism Structure
O2 complete loss of activity upon purification of enzyme in aerobic conditions or exposure to oxygen overnight Bacillus subtilis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.36
-
dihydroxyacetone phosphate 25°C Bacillus subtilis

Metals/Ions

Metals/Ions Comment Organism Structure
Iron enzyme contains a [4Fe-4S]-cluster, coordinated by residues C110, C230, C320. 3.8 mol iron per mol of enzyme Bacillus subtilis

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
41000
-
3 * 41000, calculated Bacillus subtilis
124000
-
gel filtration Bacillus subtilis

Organism

Organism UniProt Comment Textmining
Bacillus subtilis
-
isoform NadA
-

Purification (Commentary)

Purification (Comment) Organism
recombinant protein, complete loss of activity upon purification of enzyme in aerobic conditions or exposure to oxygen overnight Bacillus subtilis

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
discontinuous enzymatic assay that couples the production of iminoaspartate by NadB with the condensation between DHAP and iminoaspartate to form quinolinic acid catalyzed by NadA. The assay is linear up to 0.25 mg of NadA, 10 microg of NadB is the lowest amount suitable to measure NadA activity, and under anaerobic conditions, NadA activity becomes independent of fumarate concentration, starting from 1 mM fumarate, but decreases at concentrations higher than 2 mM fumarate, due to inhibition of NadB by fumarate Bacillus subtilis
0.027
-
25°C, presence of oxygen to reoxidize NadB in the coupled assay Bacillus subtilis
0.05
-
25°C, presence of fumarate as electron acceptor for NadB in the coupled assay Bacillus subtilis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
dihydroxyacetone phosphate + iminoaspartate
-
Bacillus subtilis ? + H2O + phosphate
-
?

Subunits

Subunits Comment Organism
trimer 3 * 41000, calculated Bacillus subtilis

Cofactor

Cofactor Comment Organism Structure
iron-sulfur centre enzyme contains a [4Fe-4S]-cluster, coordinated by residues C110, C230, C320. 3.8 mol iron per mol of enzyme, 3.3 mol sulfur per mol of enzyme Bacillus subtilis