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Literature summary for 1.14.13.135 extracted from

  • Deveryshetty, J.; Phale, P.S.
    Biodegradation of phenanthrene by Alcaligenes sp. strain PPH: partial purification and characterization of 1-hydroxy-2-naphthoic acid hydroxylase (2010), FEMS Microbiol. Lett., 311, 93-101.
    View publication on PubMed

General Stability

General Stability Organism
1-hydroxy-2-naphthoic acid hydroxylase in the cell-free extract is stabilized by 0.1 mM 1-hydroxy-2-naphthoic acid, 5 mM FAD, 2 mM DTT, and 5% glycerol Alcaligenes sp.
repeated freezing and thawing led to inactivation of the enzyme Alcaligenes sp.

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0716
-
1-hydroxy-2-naphthoate pH 7.5, 30┬░C, with NADH Alcaligenes sp.
0.0755
-
1-hydroxy-2-naphthoate pH 7.5, 30┬░C, with NADPH Alcaligenes sp.
0.0846
-
NADPH pH 7.5, 30┬░C Alcaligenes sp.
0.087
-
NADH pH 7.5, 30┬░C Alcaligenes sp.

Metals/Ions

Metals/Ions Comment Organism Structure
additional information no effect by 1 mM of Fe2+, Fe3+, Mg2+, Mn2+, Ca2+, Zn2+, and Cu2+, and by metal chelators, such as EDTA, 2,2-dipyridyl and 1',10'-phenanthroline, at 1 mM Alcaligenes sp.

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
34000
-
2 * 34000, SDS-PAGE Alcaligenes sp.
66000
-
gel filtration Alcaligenes sp.

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1-hydroxy-2-naphthoate + NADH + O2 Alcaligenes sp. a flavoprotein monooxygenase specific for 1-hydroxy-2-naphthoic acid, no activity with any other hydroxynaphthoic acid analogues, o-phthalic acid, 3,4-dihydroxybenzoic acid, or salicylic acid. In the naphthalene route, 1-hydroxy-2-naphthoic acid is metabolized via 1,2-dihydroxynaphthalene and salicylic acid to catechol by a series of enzymatic steps similar to naphthalene metabolic pathway 1,2-dihydroxynaphthalene + NAD+ + H2O + CO2
-
?
1-hydroxy-2-naphthoate + NADPH + O2 Alcaligenes sp. a flavoprotein monooxygenase specific for 1-hydroxy-2-naphthoic acid, no activity with any other hydroxynaphthoic acid analogues, o-phthalic acid, 3,4-dihydroxybenzoic acid, or salicylic acid 1,2-dihydroxynaphthalene + NADP+ + H2O + CO2
-
?

Organism

Organism UniProt Comment Textmining
Alcaligenes sp.
-
phenanthrene-degrading
-
no activity in Pseudomonas putida
-
-
-
no activity in Pseudomonas putida CSV86
-
-
-

Purification (Commentary)

Purification (Comment) Organism
native enzyme 81fold by heat treatment at 60┬░C, ammonium sulfate fractionation and anion exchange chromatography. Additional purification steps, such as hydrophobic interaction chromatography or gel filtration, lead to the total or a significant, 70%, loss of activity, respectively, without achieving any further purification Alcaligenes sp.

Source Tissue

Source Tissue Comment Organism Textmining
culture condition:phenanthrene-grown cell
-
Alcaligenes sp.
-
culture condition:salicylate-grown cell significantly lower activity compared to phenanthrene-grown cells Alcaligenes sp.
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [┬Ámol/min/mg] Specific Activity Maximum [┬Ámol/min/mg] Comment Organism
0.0031
-
salicylate-grown cells, pH 7.5, 30┬░C Alcaligenes sp.
0.0877
-
phenanthrene-grown cells, pH 7.5, 30┬░C Alcaligenes sp.
25.3
-
purified enzyme, cofactor NADPH, pH 7.5, 30┬░C Alcaligenes sp.

Storage Stability

Storage Stability Organism
-20┬░C, purified enzyme, stabilized by 0.1 mM 1-hydroxy-2-naphthoic acid, 5 mM FAD, 2 mM DTT, and 5% glycerol, retains 100% activity, after 60 days Alcaligenes sp.

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1-hydroxy-2-naphthoate + NADH + O2 a flavoprotein monooxygenase specific for 1-hydroxy-2-naphthoic acid, no activity with any other hydroxynaphthoic acid analogues, o-phthalic acid, 3,4-dihydroxybenzoic acid, or salicylic acid Alcaligenes sp. 1,2-dihydroxynaphthalene + NAD+ + H2O + CO2
-
?
1-hydroxy-2-naphthoate + NADH + O2 a flavoprotein monooxygenase specific for 1-hydroxy-2-naphthoic acid, no activity with any other hydroxynaphthoic acid analogues, o-phthalic acid, 3,4-dihydroxybenzoic acid, or salicylic acid. In the naphthalene route, 1-hydroxy-2-naphthoic acid is metabolized via 1,2-dihydroxynaphthalene and salicylic acid to catechol by a series of enzymatic steps similar to naphthalene metabolic pathway Alcaligenes sp. 1,2-dihydroxynaphthalene + NAD+ + H2O + CO2
-
?
1-hydroxy-2-naphthoate + NADPH + O2 a flavoprotein monooxygenase specific for 1-hydroxy-2-naphthoic acid, no activity with any other hydroxynaphthoic acid analogues, o-phthalic acid, 3,4-dihydroxybenzoic acid, or salicylic acid Alcaligenes sp. 1,2-dihydroxynaphthalene + NADP+ + H2O + CO2
-
?
additional information substrate specificity, overview. No activity on salicylic acid, 3-hydroxy-2-naphthoic acid, 2-hydroxy-1-naphthoic acid, 3-hydroxybenzoic acid, 4-hydroxybenzoic acid, 3,4-dihydroxybenzoic acid, 1-naphthol, 2-naphthol, 1-naphthoic acid, 2-naphthoic acid, gentisic acid or catechol Alcaligenes sp. ?
-
?

Subunits

Subunits Comment Organism
homodimer 2 * 34000, SDS-PAGE Alcaligenes sp.

Synonyms

Synonyms Comment Organism
1-hydroxy-2-naphthoic acid hydroxylase
-
Alcaligenes sp.

Temperature Optimum [┬░C]

Temperature Optimum [┬░C] Temperature Optimum Maximum [┬░C] Comment Organism
30
-
assay at Alcaligenes sp.

Temperature Stability [┬░C]

Temperature Stability Minimum [┬░C] Temperature Stability Maximum [┬░C] Comment Organism
60
-
5 min, cell-free extract, the enzyme is stable in presence of 1-hydroxy-2-naphthoic acid Alcaligenes sp.

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
-
Alcaligenes sp.

Cofactor

Cofactor Comment Organism Structure
FAD holoenzyme contains FAD. Km is 0.0047 mM. Preparation of the apoenzyme by the acid-ammonium sulfate, at 2 M, dialysis method, the apoenzyme is colorless, inactive and loses the characteristic flavin absorption spectra but regains about 90% of maximal activity when reconstituted with FAD Alcaligenes sp.
additional information FMN is a poor substitute for FAD Alcaligenes sp.
NADH Vmax/Km is similar for NADPH and NADH Alcaligenes sp.
NADPH Vmax/Km is similar for NADPH and NADH Alcaligenes sp.

Expression

Organism Comment Expression
Alcaligenes sp. the enzyme is induced by growth on phenanthrene up

General Information

General Information Comment Organism
metabolism in the naphthalene route, 1-hydroxy-2-naphthoic acid is metabolized via 1,2-dihydroxynaphthalene and salicylic acid to catechol by a series of enzymatic steps similar to naphthalene metabolic pathway Alcaligenes sp.