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Literature summary for 1.3.1.6 extracted from

  • Pankhurst, K.L.; Mowat, C.G.; Miles, C.S.; Leys, D.; Walkinshaw, M.D.; Reid, G.A.; Chapman, S.K.
    Role of His505 in the soluble fumarate reductase from Shewanella frigidimarina (2002), Biochemistry, 41, 8551-8556.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
gene fccA, expression of wild-type and mutant flavocytochrome enzymes in the enzyme-deficient strain DELTAfccA Shewanella frigidimarina

Crystallization (Commentary)

Crystallization (Comment) Organism
mutant enzyme H505A and H505Y, hanging drop vapour diffusion method, protein solution: 6 mg/ml, 10 mM Tris-HCl, pH 8.5, well solution: 100 mM Tris-HCl, pH 7.4-8.2, 25°C, 80 mM NaCl, 16-19% PEG 8000, 10 mM fumarate, equal volume of 0.002 ml of protein solution and well solution, 10 days, cryoprotectant solution: 100 mM sodium acetate, pH 6.5, 20% PEG 8000, 10 mM fumarate, 80 mM NaCl, and 23% glycerol, X-ray diffraction structure determination and analysis at 1.8 A and 2.0 A resolution, respectively, molecular replacement Shewanella frigidimarina

Protein Variants

Protein Variants Comment Organism
H505A site-directed mutagenesis, altered binding structure of the sodium ion, crystal structure analysis, at pH 8.5 the activity of the mutant enzyme is similar to the wild-type enzyme, at pH 6.0 the ctivity is 20fold reduced Shewanella frigidimarina
H505Y site-directed mutagenesis, altered binding structure of the sodium ion, crystal structure analysis, at pH 8.5 the activity of the mutant enzyme is similar to the wild-type enzyme Shewanella frigidimarina

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.007
-
fumarate wild-type enzyme, pH 9.0, 25°C Shewanella frigidimarina
0.009
-
fumarate mutant H505A, pH 9.0, 25°C Shewanella frigidimarina
0.017
-
fumarate mutant H505Y, pH 7.5, 25°C Shewanella frigidimarina
0.021
-
fumarate mutant H505Y, pH 9.0, 25°C Shewanella frigidimarina
0.022
-
fumarate mutant H505Y, pH 6.0, 25°C Shewanella frigidimarina
0.025
-
fumarate wild-type enzyme and mutant H505Y, pH 7.2, 25°C Shewanella frigidimarina
0.028
-
fumarate wild-type enzyme, pH 7.5, 25°C Shewanella frigidimarina
0.043
-
fumarate wild-type enzyme and mutant H505A, pH 6.0, 25°C Shewanella frigidimarina
0.109
-
fumarate mutant H505A, pH 7.2, 25°C Shewanella frigidimarina
0.129
-
fumarate mutant H505A, pH 7.5, 25°C Shewanella frigidimarina

Localization

Localization Comment Organism GeneOntology No. Textmining
soluble
-
Shewanella frigidimarina
-
-

Metals/Ions

Metals/Ions Comment Organism Structure
Na+ internally bound, coordinated by 1 solvent water molecule and the backbone carbonyl oxygens of residues Thr506, Met507, Gly508, Glu534, and Thr536, structure analysis, His505 is not essential for maintaining the sodium binding site Shewanella frigidimarina

Organism

Organism UniProt Comment Textmining
Shewanella frigidimarina
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant flavocytochrome enzymes from the enzyme-deficient strain DELTAfccA Shewanella frigidimarina

Reaction

Reaction Comment Organism Reaction ID
succinate + NAD+ = fumarate + NADH + H+ His505 influences the active site Shewanella frigidimarina

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
fumarate + NADH + H+
-
Shewanella frigidimarina succinate + NAD+
-
?

Synonyms

Synonyms Comment Organism
fumarate reductase
-
Shewanella frigidimarina

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Shewanella frigidimarina

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
32
-
fumarate mutant H505A, pH 6.0, 25°C Shewanella frigidimarina
79
-
fumarate mutant H505A, pH 7.2, 25°C Shewanella frigidimarina
101
-
fumarate mutant H505A, pH 7.5, 25°C Shewanella frigidimarina
105
-
fumarate mutant H505A, pH 9.0, 25°C Shewanella frigidimarina
210
-
fumarate wild-type enzyme, pH 9.0, 25°C Shewanella frigidimarina
240
-
fumarate mutant H505Y, pH 9.0, 25°C Shewanella frigidimarina
354
-
fumarate mutant H505Y, pH 7.2, 25°C Shewanella frigidimarina
363
-
fumarate mutant H505Y, pH 7.5, 25°C Shewanella frigidimarina
370
-
fumarate wild-type enzyme, pH 7.5, 25°C Shewanella frigidimarina
377
-
fumarate mutant H505Y, pH 6.0, 25°C Shewanella frigidimarina
509
-
fumarate wild-type enzyme, pH 7.2, 25°C Shewanella frigidimarina
658
-
fumarate wild-type enzyme, pH 6.0, 25°C Shewanella frigidimarina

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
additional information
-
-
Shewanella frigidimarina

pH Range

pH Minimum pH Maximum Comment Organism
6 9
-
Shewanella frigidimarina

Cofactor

Cofactor Comment Organism Structure
flavocytochrome c3
-
Shewanella frigidimarina
NADH
-
Shewanella frigidimarina