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Literature summary for 1.7.1.3 extracted from

  • Minagawa, N.; Yoshimoto, A.
    Purification and characterization of the assimilatory NADPH-nitrate reductase of Aspergillus nidulans (1982), J. Biochem., 91, 761-774.
    View publication on PubMed

Activating Compound

Activating Compound Comment Organism Structure
FAD activates 10fold by addition of 0.004 mM, stabilizes against heat inactivation Aspergillus nidulans

Inhibitors

Inhibitors Comment Organism Structure
azide
-
Aspergillus nidulans
cyanide 99% inhibition at 10 mM Aspergillus nidulans
p-chloromercuribenzoate reversion of the inhibition by addition of reduced glutathione Aspergillus nidulans

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.045
-
NADPH + nitrate Aspergillus nidulans
0.2
-
nitrate cosubstrate NADPH Aspergillus nidulans
2.5
-
FADH2 + nitrate Aspergillus nidulans
3
-
FMNH2 + nitrate Aspergillus nidulans

Metals/Ions

Metals/Ions Comment Organism Structure
Molybdenum molybdoflavoprotein Aspergillus nidulans

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
38000
-
2 * 59000 + 2 * 38000, SDS-PAGE, after heat-treatment all polypeptide chains are 59 kDa Aspergillus nidulans
59000
-
2 * 59000 + 2 * 38000, SDS-PAGE, after heat-treatment all polypeptide chains are 59 kDa Aspergillus nidulans
180000
-
gel filtration, sucrose density gradient Aspergillus nidulans

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
nitrate + NADPH Aspergillus nidulans
-
nitrite + NADP+ + H2O
-
?

Organism

Organism UniProt Comment Textmining
Aspergillus nidulans
-
IFO No. 6083
-

Purification (Commentary)

Purification (Comment) Organism
ammonium slufate precipitation, ion-exchange, gel filtration, hydroxylapatite column. FAD and EDTA essential in all buffers during purification Aspergillus nidulans

Specific Activity [micromol/min/mg]

Specific Activity Minimum [┬Ámol/min/mg] Specific Activity Maximum [┬Ámol/min/mg] Comment Organism
15.5
-
-
Aspergillus nidulans

Storage Stability

Storage Stability Organism
-80┬░C, 0.02 mM FAD, several months with only slight loss of the activity Aspergillus nidulans

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
nitrate + NADPH
-
Aspergillus nidulans nitrite + NADP+ + H2O
-
?
nitrate + NADPH 4 activities: NADPH-nitrate reductase, FADH-nitrate reductase, reduced methyl viologen-nitrate reductase and NADPH-cytochrome c reductase Aspergillus nidulans nitrite + NADP+ + H2O
-
?
nitrate + NADPH catalyzes the NADPH-linked reduction of ferricyanide and 2,6-dichlorophenolindophenol, chlorate- and bromate-dependent NADPH oxidation, and FMNH-linked nitrate reduction Aspergillus nidulans nitrite + NADP+ + H2O
-
?

Subunits

Subunits Comment Organism
tetramer 2 * 59000 + 2 * 38000, SDS-PAGE, after heat-treatment all polypeptide chains are 59 kDa Aspergillus nidulans

Temperature Optimum [┬░C]

Temperature Optimum [┬░C] Temperature Optimum Maximum [┬░C] Comment Organism
30
-
assay at Aspergillus nidulans

Temperature Stability [┬░C]

Temperature Stability Minimum [┬░C] Temperature Stability Maximum [┬░C] Comment Organism
37
-
absence of FAD, rapid loss of activity, decreases to 50% in less than 20 min Aspergillus nidulans

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7 8
-
Aspergillus nidulans
7.4
-
assay at Aspergillus nidulans

Cofactor

Cofactor Comment Organism Structure
cytochrome b557 involved in intracellular electron transport from NADPH to nitrate Aspergillus nidulans
FAD contains 5.91-7.78 nmol FAD per mg of protein, activation of NADPH-nitrate reductase activity Aspergillus nidulans
FAD molybdoflavoprotein Aspergillus nidulans
additional information NADH: no effect, presence of protoheme IX Aspergillus nidulans