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Literature summary for 1.1.2.3 extracted from

  • Cenas, N.; Le, K.H.; Terrier, M.; Lederer, F.
    Potentiometric and further kinetic characterization of the flavin-binding domain of Saccharomyces cerevisiae flavocytochrome b2. Inhibition by anions binding in the active site (2007), Biochemistry, 46, 4661-4670.
    View publication on PubMed
Search for more literature for 1.1.2.3

Cloned(Commentary)

Cloned (Comment) Organism
recombinant wild-type Fcb2 and its recombinant FMN-binding domain (FDH domain) are expressed in Escherichia coli Saccharomyces cerevisiae

Protein Variants

Protein Variants Comment Organism
R289K kcat (1/sec) (substrate: L-lactate): 8.6 (in 200 mM phosphate: 9.2, in 400 mM potassium acetate: 8.8, in 400 mM KCl: 9.2, in 400 mM KBr: 7.8), Km (mM) (substrate: L-lactate): 7.0 (in 200 mM phosphate: 8.7, in 400 mM potassium acetate: 9.2, in 400 mM KCl: 6.5, in 400 mM KBr: 5.8). Mutant is not sensitive for excess lactate concentration. In contrast to the wild-type enzyme high concentrations of acetate, phosphate, chloride and bromide show no influence on the mutant enzyme Saccharomyces cerevisiae

Inhibitors

Inhibitors Comment Organism Structure
acetate
-
 Saccharomyces cerevisiae
bromide
-
 Saccharomyces cerevisiae
chloride
-
 Saccharomyces cerevisiae
additional information enzyme is inhibited by high substrate concentrations Saccharomyces cerevisiae
phosphate
-
 Saccharomyces cerevisiae
pyruvate
-
 Saccharomyces cerevisiae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.66
-
 L-lactate FDH domain, reaction conditions: 13 mM ferricyanide, 100 mM phosphate buffer, 1 mM EDTA, pH 7, 30°C Saccharomyces cerevisiae
0.66
-
 L-lactate FDH domain, reaction conditions: 13 mM ferricyanide, 100 mM phosphate buffer, 1 mM EDTA, pH 7.5, 25°C Saccharomyces cerevisiae
0.86
-
 L-lactate FDH domain, reaction conditions: 13 mM ferricyanide, 10 mM Tris/HCl buffer, 0.1 M NaCl, pH 7.5, 25°C Saccharomyces cerevisiae
0.89
-
 L-lactate stopped-flow kinetic parameters for flavin reduction by L-lactate using holo-enzyme, reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, in the absence of ferrocyanide Saccharomyces cerevisiae
0.9
-
 L-lactate influence of anions (200 mM phsophate) on the wild-type steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 1.5 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
0.94
-
 L-lactate influence of anions (400 mM KBr) on the wild-type steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 1.5 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
1.03
-
 L-lactate stopped-flow kinetic parameters for flavin reduction by L-lactate using FDH domain, reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, in the absence of ferrocyanide Saccharomyces cerevisiae
1.18
-
 L-lactate FDH domain, reaction conditions: 13 mM ferricyanide, 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C Saccharomyces cerevisiae
1.5
-
 L-lactate influence of anions (400 mM KCl) on the wild-type steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 1.5 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
2.3
-
 L-lactate influence of anions (400 mM KCl) on the FDH domain steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, constant 10 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
2.7
-
 L-lactate influence of anions (400 mM potassium acetate) on the wild-type steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 1.5 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
2.8
-
 L-lactate influence of anions (300 mM phsophate) on the FDH domain steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, constant 10 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
4
-
 L-lactate influence of anions (400 mM KBr) on the FDH domain steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, constant 10 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
4.6
-
 L-lactate influence of anions (400 mM potassium acetate) on the FDH domain steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, constant 10 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
5.8
-
 L-lactate influence of anions (400 mM KBr) on mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
6.5
-
 L-lactate influence of anions (400 mM KCl) on mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
7
-
 L-lactate mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
8.7
-
 L-lactate influence of anions (200 mM phsophate) on mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
9.2
-
 L-lactate influence of anions (400 mM potassium acetate) on mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae

Organism

Organism UniProt Comment Textmining
Saccharomyces cerevisiae
-
-
-

Storage Stability

Storage Stability Organism
-70°C in 0.1 M phosphate buffer, 1 mM EDTA, pH 7, 10 mM DL-lactate is added to the Fcb2 preparations to keep the enzyme in the reduced state Saccharomyces cerevisiae

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
L-lactate + ferricyanide
-
 Saccharomyces cerevisiae pyruvate + ferrocyanide + H+
-
 ?

Synonyms

Synonyms Comment Organism
cytochrome c oxido reductase
-
 Saccharomyces cerevisiae
flavocytochrome b2
-
 Saccharomyces cerevisiae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
7.8
-
 L-lactate influence of anions (400 mM KBr) on mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
8.6
-
 L-lactate mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
8.8
-
 L-lactate influence of anions (400 mM potassium acetate) on mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
9.2
-
 L-lactate influence of anions (200 mM phsophate) on mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
9.2
-
 L-lactate influence of anions (400 mM KCl) on mutant R289K steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 2 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
45
-
 L-lactate influence of anions (400 mM KBr) on the FDH domain steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, constant 10 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
60
-
 L-lactate influence of anions (400 mM KCl) on the FDH domain steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, constant 10 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
61
-
 L-lactate influence of anions (400 mM KBr) on the wild-type steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 1.5 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
71
-
 L-lactate influence of anions (200 mM phsophate) on the wild-type steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 1.5 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
75
-
 L-lactate influence of anions (400 mM KCl) on the wild-type steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 1.5 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
86
-
 L-lactate influence of anions (400 mM potassium acetate) on the FDH domain steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, constant 10 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
101
-
 L-lactate influence of anions (300 mM phsophate) on the FDH domain steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, constant 10 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
113
-
 L-lactate influence of anions (400 mM potassium acetate) on the wild-type steady-state kinetic. Reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, 1.5 mM ferricyanide and variable L-lactate concentration Saccharomyces cerevisiae
117
-
 L-lactate holo-enzyme, reaction conditions: 1 or 2 mM ferricyanide, 100 mM phosphate buffer, 1mM EDTA, pH 7, 5°C Saccharomyces cerevisiae
133
-
 L-lactate FDH domain, reaction conditions: 13 mM ferricyanide, 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C Saccharomyces cerevisiae
144
-
 L-lactate stopped-flow kinetic parameters for flavin reduction by L-lactate using holo-enzyme, reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, in the absence of ferrocyanide Saccharomyces cerevisiae
149
-
 L-lactate stopped-flow kinetic parameters for flavin reduction by L-lactate using FDH domain, reaction conditions: 100 mM phosphate buffer, 1 mM EDTA, pH 7, 5°C, in the absence of ferrocyanide Saccharomyces cerevisiae
214
-
 L-lactate FDH domain, reaction conditions: 13 mM ferricyanide, 10 mM Tris/HCl buffer, 0.1 M NaCl, pH 7.5, 25°C Saccharomyces cerevisiae
240
-
 L-lactate FDH domain, reaction conditions: 13 mM ferricyanide, 100 mM phosphate buffer, 1 mM EDTA, pH 7.5, 25°C Saccharomyces cerevisiae
259
-
 L-lactate FDH domain, reaction conditions: 13 mM ferricyanide, 100 mM phosphate buffer, 1 mM EDTA, pH 7, 30°C Saccharomyces cerevisiae

Cofactor

Cofactor Comment Organism Structure
FMN
-
 Saccharomyces cerevisiae

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
6
-
 pyruvate using the FDH domain at varied ferrocyanide concentrations and a fixed L-lactate concentration pyruvate behaves as a mixed-type inhibitor toward ferrocyanide Saccharomyces cerevisiae
6
-
 pyruvate using the FDH domain at varied L-lactate concentrations and a fixed ferrocyanide concentration pyruvate behaves as a mixed-type inhibitor toward L-lactate Saccharomyces cerevisiae
9.7
-
 pyruvate in pre-steady-state kinetic studies of the FDH domain reduction and reoxidation, pyruvate acts as a competitive inhibitor of L-lactate Saccharomyces cerevisiae
170
-
 acetate in pre-steady-state kinetic studies of the FDH domain reduction and reoxidation, acetate acts as a competitive inhibitor of L-lactate Saccharomyces cerevisiae
450
-
 chloride in pre-steady-state kinetic studies of the FDH domain reduction and reoxidation, chloride acts as a competitive inhibitor of L-lactate Saccharomyces cerevisiae