Cloned (Comment) | Organism |
---|---|
recombinant expression of wild-type and mutant enzymes | Saccharomyces cerevisiae |
Crystallization (Comment) | Organism |
---|---|
purified recombinant enzyme mutant N195A, sitting drop vapour diffusion method, mixing of protein solution, containing 25 mM HEPES, 25 mM NaCl, and 2% glycerol, pH 7.5, in a 1:1 ratio with buffer containing 30 mM diethylene glycol, 30 mM triethylene glycol, 30 mM tetraethylene glycol, 30 mM pentaethylene glycol, 10% ethylene glycol, 20% PEG 8000, and 0.1 MES-imidazole, pH 6.5, 1 week, X-ray diffraction structure determination and analysis at 2.0 A resolution | Saccharomyces cerevisiae |
Protein Variants | Comment | Organism |
---|---|---|
D94N | site-directed mutagenesis, the mutation primarily affects the reductive half-reaction. The mutant shows 20-40fold reduced rate constant for flavin reduction with spermine and 450fold with N1-acetylspermine compared to the wild-type enzyme | Saccharomyces cerevisiae |
N195A | site-directed mutagenesis, the mutation primarily affects the reductive half-reaction. The mutant shows 20-40fold reduced rate constant for flavin reduction with spermine and 450fold with N1-acetylspermine compared to the wild-type enzyme. Mutant N195A enzyme shows structure with a molecule of tetraethylene glycol in the active site, the mutation has no effect on the protein structure | Saccharomyces cerevisiae |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | steady-state kinetics and rapid-reaction kinetics of wild-type and mutant enzymes, overview | Saccharomyces cerevisiae | |
0.011 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant wild-type Fms1 | Saccharomyces cerevisiae | |
0.015 | - |
O2 | pH 9.0, 25°C, recombinant mutant N195A, with spermine | Saccharomyces cerevisiae | |
0.02 | - |
O2 | pH 9.0, 25°C, recombinant mutant N195A, with N1-acetylspermine | Saccharomyces cerevisiae | |
0.03 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant mutant D94N | Saccharomyces cerevisiae | |
0.0436 | - |
O2 | pH 9.0, 25°C, recombinant wild-type Fms1, with N1-acetylspermine | Saccharomyces cerevisiae | |
0.091 | - |
O2 | pH 9.0, 25°C, recombinant wild-type Fms1, with spermine | Saccharomyces cerevisiae | |
0.097 | - |
O2 | pH 9.0, 25°C, recombinant mutant D94N, with N1-acetylspermine | Saccharomyces cerevisiae | |
0.104 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant mutant N195A | Saccharomyces cerevisiae | |
0.118 | - |
spermine | pH 9.35, 25°C, recombinant wild-type Fms1 | Saccharomyces cerevisiae | |
0.127 | - |
spermine | pH 9.35, 25°C, recombinant mutant N195A | Saccharomyces cerevisiae | |
0.192 | - |
O2 | pH 9.0, 25°C, recombinant mutant D94N, with spermine | Saccharomyces cerevisiae | |
0.32 | - |
spermine | pH 9.35, 25°C, recombinant mutant D94N | Saccharomyces cerevisiae |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
N1-acetylspermine + O2 + H2O | Saccharomyces cerevisiae | - |
spermidine + N-acetyl-3-aminopropanal + H2O2 | - |
? | |
spermine + O2 + H2O | Saccharomyces cerevisiae | - |
spermidine + 3-aminopropanal + H2O2 | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Saccharomyces cerevisiae | P50264 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and mutant enzymes | Saccharomyces cerevisiae |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
N1-acetylspermine + O2 + H2O | - |
Saccharomyces cerevisiae | spermidine + N-acetyl-3-aminopropanal + H2O2 | - |
? | |
spermine + O2 + H2O | - |
Saccharomyces cerevisiae | spermidine + 3-aminopropanal + H2O2 | - |
? |
Synonyms | Comment | Organism |
---|---|---|
Fms1 | - |
Saccharomyces cerevisiae |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Saccharomyces cerevisiae |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
2 | - |
spermine | pH 9.35, 25°C, recombinant mutant D94N | Saccharomyces cerevisiae | |
4.9 | - |
spermine | pH 9.35, 25°C, recombinant mutant N195A | Saccharomyces cerevisiae | |
8.1 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant mutant N195A | Saccharomyces cerevisiae | |
9.7 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant mutant D94N | Saccharomyces cerevisiae | |
15.1 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant wild-type Fms1 | Saccharomyces cerevisiae | |
39 | - |
spermine | pH 9.35, 25°C, recombinant wild-type Fms1 | Saccharomyces cerevisiae |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
9 | - |
substrate N1-acetylspermine | Saccharomyces cerevisiae |
9.35 | - |
substrate spermine | Saccharomyces cerevisiae |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
FAD | dependent on | Saccharomyces cerevisiae |
General Information | Comment | Organism |
---|---|---|
additional information | the active site of Fms1 contains three amino acid residues positioned to interact with the polyamine substrate, His67, Asn195, and Asp94. These three residues form a hydrogen-bonding triad with Asn195 being the central residue. His67 is important both for interacting with the substrate and for maintaining the hydrogen bonds in the triad | Saccharomyces cerevisiae |
physiological function | flavoprotein Fms1 catalyzes the oxidation of spermine in the biosynthetic pathway for pantothenic acid | Saccharomyces cerevisiae |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
4.1 | - |
spermine | pH 9.35, 25°C, recombinant mutant D94N | Saccharomyces cerevisiae | |
38.5 | - |
spermine | pH 9.35, 25°C, recombinant mutant N195A | Saccharomyces cerevisiae | |
78 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant mutant N195A | Saccharomyces cerevisiae | |
100 | - |
O2 | pH 9.0, 25°C, recombinant mutant D94N, with N1-acetylspermine | Saccharomyces cerevisiae | |
104 | - |
O2 | pH 9.0, 25°C, recombinant mutant D94N, with spermine | Saccharomyces cerevisiae | |
320 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant mutant D94N | Saccharomyces cerevisiae | |
327 | - |
O2 | pH 9.0, 25°C, recombinant mutant N195A, with spermine | Saccharomyces cerevisiae | |
330 | - |
spermine | pH 9.35, 25°C, recombinant wild-type Fms1 | Saccharomyces cerevisiae | |
358 | - |
O2 | pH 9.0, 25°C, recombinant wild-type Fms1, with N1-acetylspermine | Saccharomyces cerevisiae | |
405 | - |
O2 | pH 9.0, 25°C, recombinant mutant N195A, with N1-acetylspermine | Saccharomyces cerevisiae | |
428 | - |
O2 | pH 9.0, 25°C, recombinant wild-type Fms1, with spermine | Saccharomyces cerevisiae | |
1400 | - |
N1-acetylspermine | pH 9.0, 25°C, recombinant wild-type Fms1 | Saccharomyces cerevisiae |