1.13.11.6: 3-hydroxyanthranilate 3,4-dioxygenase

This is an abbreviated version, for detailed information about 3-hydroxyanthranilate 3,4-dioxygenase, go to the full flat file.

Reaction

3-Hydroxyanthranilate
+
O2
=
2-amino-3-carboxymuconate semialdehyde

Synonyms

3-HAD, 3-HAO, 3-hydroxyanthranilate 3,4-dioxygenase, 3-hydroxyanthranilate oxygenase, 3-hydroxyanthranilic acid oxidase, 3-hydroxyanthranilic acid oxygenase, 3-hydroxyanthranilic oxygenase, 3HAO, EC 1.13.1.6, HAD, oxygenase, 3-hydroxyanthranilate 3,4-di-

ECTree

     1 Oxidoreductases
         1.13 Acting on single donors with incorporation of molecular oxygen (oxygenases)
             1.13.11 With incorporation of two atoms of oxygen
                1.13.11.6 3-hydroxyanthranilate 3,4-dioxygenase

Purification

Purification on EC 1.13.11.6 - 3-hydroxyanthranilate 3,4-dioxygenase

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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
bovine kidney homogenized in 0.1 M potassium dihydrogen phosphate buffer with 20% glycerol and 3 mM 2-mercaptoethanol, pH 7.4, centrifuged, protein fraction of supernatant that precipitates from 34-62% saturated (NH4)2SO4 is resuspended in 5 mM potassium dihydrogen phosphate buffer with 20% glycerol and 3 mM 2-mercaptoethanol, pH 7.4, dialyzed against same buffer containing protease inhibitor PMSF (0.1 mM), applied to DEAE Sephadex A-50 column, fractions with enzyme activity pooled and concentrated by precipitation with 80%-saturated (NH4)2SO4, dialyzed against 10 mM Tris-HCl with 20% glycerol and 3 mM 2-mercaptoethanol, pH 7.4, applied to Blue-Sepharose CL-4B column, concentration of active fractions by ultrafiltration through YM-10 membrane
pure liver enzyme, partielly purified brain enzyme
-
recombinant His6-MBP-tagged enzyme from Escherichia coli by nickel affinity chromatography and cleavage of the His-tag by TEV protease, elimination of the tags by nickel affinity and amylose affinity chromatography, te final step is gel filtration
-
succesive size exclusion and affinity columns
-