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Literature summary extracted from

  • Li, H.; Xie, K.; Huang, H.; Wang, S.
    6-hydroxy-3-succinoylpyridine hydroxylase catalyzes a central step of nicotine degradation in Agrobacterium tumefaciens S33 (2014), PLoS ONE, 9, e103324 .
    View publication on PubMedView publication on EuropePMC

Application

EC Number Application Comment Organism
1.14.13.163 synthesis development of an efficient process to transform HSP into 2,5-dihydroxypyridine (2,5-DHP) with heterologously expressed HSP hydroxylase and NADH-regenerating system, because 2,5-DHP, the product of the reaction catalyzed by HSP hydroxylase, is a valuable precursor for the chemical synthesis of 5-aminolevulinic acid, which is applied as a plant growth hormone, a herbicide and in cancer therapy Agrobacterium tumefaciens

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.14.13.163 DNA and amino acid sequence determination and analysis, sequence comparisons, RT-PCR expression analysis, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3) Agrobacterium tumefaciens

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.14.13.163 90000
-
 gel filtration Agrobacterium tumefaciens

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.14.13.163 4-(6-hydroxypyridin-3-yl)-4-oxobutanoate + 2 NADH + 2 H+ + O2 Agrobacterium tumefaciens
-
 2,5-dihydroxypyridine + succinate semialdehyde + 2 NAD+ + H2O
-
 ?
1.14.13.163 4-(6-hydroxypyridin-3-yl)-4-oxobutanoate + 2 NADH + 2 H+ + O2 Agrobacterium tumefaciens S33
-
 2,5-dihydroxypyridine + succinate semialdehyde + 2 NAD+ + H2O
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.14.13.163 Agrobacterium tumefaciens W8E0Y0 i.e. Rhizobium radiobacter
-
1.14.13.163 Agrobacterium tumefaciens S33 W8E0Y0 i.e. Rhizobium radiobacter
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.14.13.163 native enzyme 64.4fold from Agrobacterium tumefaciens strain S33 cells by ammonium sulfate fractionation, hydrophobic interaction chromatography, desalting gel filtration, and two different steps of anion exchange chromatography, followed by gel filtration. Recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, anion exchange chromatography, and ultrafiltration. During purification, all the buffers are supplemented with 0.005 mM FAD. When FAD is omitted, the purification yield is very low Agrobacterium tumefaciens

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.14.13.163 10.3
-
 purified native enzyme, pH 8.0, 30°C Agrobacterium tumefaciens
1.14.13.163 18.8
-
 purified recombinant enzyme, pH 8.0, 30°C Agrobacterium tumefaciens

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.14.13.163 4-(6-hydroxypyridin-3-yl)-4-oxobutanoate + 2 NADH + 2 H+ + O2
-
 Agrobacterium tumefaciens 2,5-dihydroxypyridine + succinate semialdehyde + 2 NAD+ + H2O
-
 ?
1.14.13.163 4-(6-hydroxypyridin-3-yl)-4-oxobutanoate + 2 NADH + 2 H+ + O2 oxidative decarboxylation of HSP to 2,5-dihydroxypyridine in presence of NADH and FAD Agrobacterium tumefaciens 2,5-dihydroxypyridine + succinate semialdehyde + 2 NAD+ + H2O
-
 ?
1.14.13.163 4-(6-hydroxypyridin-3-yl)-4-oxobutanoate + 2 NADH + 2 H+ + O2
-
 Agrobacterium tumefaciens S33 2,5-dihydroxypyridine + succinate semialdehyde + 2 NAD+ + H2O
-
 ?
1.14.13.163 4-(6-hydroxypyridin-3-yl)-4-oxobutanoate + 2 NADH + 2 H+ + O2 oxidative decarboxylation of HSP to 2,5-dihydroxypyridine in presence of NADH and FAD Agrobacterium tumefaciens S33 2,5-dihydroxypyridine + succinate semialdehyde + 2 NAD+ + H2O
-
 ?
1.14.13.163 additional information no activity with p-nitrophenol or 6-hydroxynicotinate as substrates Agrobacterium tumefaciens ?
-
 ?
1.14.13.163 additional information no activity with p-nitrophenol or 6-hydroxynicotinate as substrates Agrobacterium tumefaciens S33 ?
-
 ?

Subunits

EC Number Subunits Comment Organism
1.14.13.163 homodimer 2 * 45000, SDS-PAGE Agrobacterium tumefaciens

Synonyms

EC Number Synonyms Comment Organism
1.14.13.163 6-hydroxy-3-succinoylpyridine hydroxylase
-
 Agrobacterium tumefaciens
1.14.13.163 AWN88_01205 gene name, UniProt Agrobacterium tumefaciens
1.14.13.163 HSP hydroxylase
-
 Agrobacterium tumefaciens
1.14.13.163 NADH-dependent HSP hydroxylase
-
 Agrobacterium tumefaciens

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.14.13.163 30
-
 assay at Agrobacterium tumefaciens

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.14.13.163 8
-
-
 Agrobacterium tumefaciens

Cofactor

EC Number Cofactor Comment Organism Structure
1.14.13.163 FAD required for activity Agrobacterium tumefaciens
1.14.13.163 NADH dependent on Agrobacterium tumefaciens

General Information

EC Number General Information Comment Organism
1.14.13.163 metabolism 6-hydroxy-3-succinoylpyridine hydroxylase catalyzes a central step of nicotine degradation. 6-Hydroxy-3-succinoylpyridine (HSP) is a key intermediate connecting the two pathways, pyridine pathway and pyrrolidine pathway, overview Agrobacterium tumefaciens
1.14.13.163 physiological function the key enzyme HSP hydroxylase is involved in the fused nicotine degradation pathway of the pyridine and pyrrolidine pathways Agrobacterium tumefaciens