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Literature summary for 1.5.3.17 extracted from

  • Adachi, M.; Taylor, A.; Hart, P.; Fitzpatrick, P.
    Mechanistic and structural analyses of the roles of active site residues in yeast polyamine oxidase Fms1: characterization of the N195A and D94N enzymes (2012), Biochemistry, 51, 8690-8697.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of wild-type and mutant enzymes Saccharomyces cerevisiae

Crystallization (Commentary)

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

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 [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/ Products (Substrates)

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

Organism UniProt Comment Textmining
Saccharomyces cerevisiae P50264
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes Saccharomyces cerevisiae

Substrates and Products (Substrate)

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

Synonyms Comment Organism
Fms1
-
Saccharomyces cerevisiae

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Saccharomyces cerevisiae

Turnover Number [1/s]

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

pH Optimum Minimum pH Optimum Maximum Comment Organism
9
-
substrate N1-acetylspermine Saccharomyces cerevisiae
9.35
-
substrate spermine Saccharomyces cerevisiae

Cofactor

Cofactor Comment Organism Structure
FAD dependent on Saccharomyces cerevisiae

General Information

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 [mM/s]

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