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Literature summary for 1.4.3.2 extracted from

  • Hess, M.C.; Bloess, S.; Risse, J.M.; Friehs, K.; Fischer von Mollard, G.
    Recombinant expression of an l-amino acid oxidase from the fungus Hebeloma cylindrosporum in Pichia pastoris including fermentation (2020), MicrobiologyOpen, 9, e1112 .
    View publication on PubMed
Search for more literature for 1.4.3.2

Activating Compound

Activating Compound Comment Organism Structure
additional information enzyme is activated by incubation for 10 min at pH 3 Hebeloma cylindrosporum

Application

Application Comment Organism
synthesis expression of isoform LAOO4 in the yeast Pichia pastoris with a 9His-tag and comparison with the 6His-LAAO4 expressed in Escherichia coli. The expression of the enzyme with an ER-signal sequence in Pastoris pastoris results in a glycosylated, secreted protein. The enzymatic activity without activation is higher after expression in Pichia pastoris compared to Escherichia coli. Due to treatment with acidic pH, a striking increase of activity can be detected for both expression systems resulting in similar specific activities after acid activation Hebeloma cylindrosporum

Cloned(Commentary)

Cloned (Comment) Organism
expression in Pichia pastoris Hebeloma cylindrosporum

Protein Variants

Protein Variants Comment Organism
N54A/N164A/N193A/N331A mutation of the potential glycosylatoon sites, mutant enzyme has the same mobility as the lower band of 9His-LAAO4 treated with PNGaseF Hebeloma cylindrosporum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.04
-
 L-leucine untreated recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
0.07
-
 L-phenylalanine acid-activited recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
0.07
-
 L-phenylalanine untreated recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
0.16
-
 L-methionine untreated recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
0.29
-
 L-leucine acid-activited recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
0.41
-
 L-methionine acid-activated recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
0.51
-
 L-glutamine untreated recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
0.59
-
 L-glutamine acid-activited recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
1.12
-
 L-phenylalanine methyl ester acid-activited recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
1.85
-
 L-phenylalanine methyl ester untreated recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
3.01
-
 L-leucine methyl ester untreated recombinant protein, pH 7, 30°C Hebeloma cylindrosporum
3.58
-
 L-leucine methyl ester acid-activited recombinant protein, pH 7, 30°C Hebeloma cylindrosporum

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
216000
-
 gel filtration Hebeloma cylindrosporum

Organism

Organism UniProt Comment Textmining
Hebeloma cylindrosporum S4S6Z0 isoform LAAO4
-

Posttranslational Modification

Posttranslational Modification Comment Organism
glycoprotein sequence contains four putative N-glycosylation sites predicted at amino acid residues N54, N164, N193, and N331 Hebeloma cylindrosporum

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
2
-
 untreated enzyme, expressed as 6His-LAAO4 in Escherichia coli, pH 7, 30°C Hebeloma cylindrosporum
20 40 acid-treated enzyme, expressed as 9His-LAAO4 in Pichia pastoris, as well as 6His-LAAO4 expressed in Escherichia coli, pH 7, 30°C Hebeloma cylindrosporum
1020
-
 untreated enzyme, expressed as 9His-LAAO4 in Pichia pastoris, pH 7, 30°C Hebeloma cylindrosporum

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
4-chloro-DL-phenylalanine + H2O + O2
-
 Hebeloma cylindrosporum 2-oxo-3-(4-chlorophenyl)-propanoate + NH3 + H2O2
-
 ?
DL-homophenylalanine + H2O + O2
-
 Hebeloma cylindrosporum 2-oxo-4-phenylbutanoate + NH3 + H2O2
-
 ?
L-alanine + H2O + O2
-
 Hebeloma cylindrosporum 2-oxopropanoate + NH3 + H2O2
-
 ?
L-arginine + H2O + O2
-
 Hebeloma cylindrosporum 2-oxo-5-guanidinopentanoate + NH3 + H2O2
-
 ?
L-asparagine + H2O + O2
-
 Hebeloma cylindrosporum 2-oxo-3-phenylpropanoate + NH3 + H2O2
-
 ?
L-glutamate + H2O + O2
-
 Hebeloma cylindrosporum 2-oxoglutarate + NH3 + H2O2
-
 ?
L-glutamate dimethyl ester + H2O + O2
-
 Hebeloma cylindrosporum 2-oxoglutarate dimethyl ester + NH3 + H2O2
-
 ?
L-glutamine + H2O + O2
-
 Hebeloma cylindrosporum 2-oxo-4-carbamoylbutanoic acid + NH3 + H2O2
-
 ?
L-histidine + H2O + O2
-
 Hebeloma cylindrosporum 3-(1H-imidazol-4-yl)-2-oxopropanoate + NH3 + H2O2
-
 ?
L-isoleucine + H2O + O2
-
 Hebeloma cylindrosporum 3-methyl-2-oxo-pentanoate + NH3 + H2O2
-
 ?
L-leucine + H2O + O2
-
 Hebeloma cylindrosporum 4-methyl-2-oxopentanoate + NH3 + H2O2
-
 ?
L-leucine methyl ester + H2O + O2
-
 Hebeloma cylindrosporum methyl 4-methyl-2-oxopentanoate + NH3 + H2O2
-
 ?
L-lysine + H2O + O2
-
 Hebeloma cylindrosporum 6-amino-2-oxohexanoate + NH3 + H2O2
-
 ?
L-methionine + H2O + O2
-
 Hebeloma cylindrosporum 4-methylsulfanyl-2-oxobutanoate + NH3 + H2O2
-
 ?
L-methionine methyl ester + H2O + O2
-
 Hebeloma cylindrosporum methyl 4-methylsulfanyl-2-oxobutanoate + NH3 + H2O2
-
 ?
L-norleucine + H2O + O2
-
 Hebeloma cylindrosporum 2-oxohexanoate + NH3 + H2O2
-
 ?
L-ornithine + H2O + O2
-
 Hebeloma cylindrosporum 5-amino-2-oxopentanoate + NH3 + H2O2
-
 ?
L-phenylalanine + H2O + O2
-
 Hebeloma cylindrosporum 2-oxo-3-phenylpropanoate + NH3 + H2O2
-
 ?
L-phenylalanine methyl ester + H2O + O2
-
 Hebeloma cylindrosporum methyl 2-oxo-3-phenylpropanoate + NH3 + H2O2
-
 ?
L-tyrosine + H2O + O2
-
 Hebeloma cylindrosporum 3-(4-hydroxyphenyl)pyruvate + NH3 + H2O2
-
 ?

Subunits

Subunits Comment Organism
trimer 3 * 69600, calculated from sequence, 3 * 75000, SDS-PAGE of 9His-LAAO4 expressed in Pichia pastoris Hebeloma cylindrosporum

Synonyms

Synonyms Comment Organism
LAAO4
-
 Hebeloma cylindrosporum

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6 9 broad, acid-treated enzyme Hebeloma cylindrosporum

pH Range

pH Minimum pH Maximum Comment Organism
3
-
 complete loss of activity below Hebeloma cylindrosporum

Cofactor

Cofactor Comment Organism Structure
FAD FAD loading of the enzyme expressed in Pichia pastoris is 80%-100% in different expressions Hebeloma cylindrosporum