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Literature summary for 1.2.3.4 extracted from

  • Whittaker, M.M.; Pan, H.Y.; Yukl, E.T.; Whittaker, J.W.
    Burst kinetics and redox transformations of the active site manganese ion in oxalate oxidase: Implications for the catalytic mechanism (2007), J. Biol. Chem., 282, 7011-7023.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
recombinant barley oxalate oxidase expressed in Pichia pastoris (X33) Hordeum vulgare

Protein Variants

Protein Variants Comment Organism
S49A nonglycosylated oxalate oxidase is produced by site-directed mutagenesis (S49A) Hordeum vulgare

General Stability

General Stability Organism
based on the absoption changes at 325 nm, it is possible to estimate the half-life of the Mn5+ species at room temperature: t1/2 = 42 h (pH 4) or 95 h (pH 7) Hordeum vulgare

Inhibitors

Inhibitors Comment Organism Structure
additional information the presence of either superoxide dismutase or manganese catalase in the assay mixture dramatically accelerates turnover inactivation and resultes in a vanishingly small Vs value in the steady state Hordeum vulgare

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.78
-
oxalate oxalate oxidase activity is measured by oxygen uptake assay with a Clark oxygen electrode in a thermostated cell (25°C). The Km evalutated from initial velocity data exhibits a strong pH dependence with limiting slopes (versus pH) of 0.9 (below pH 4) and 1.5 (above pH 4) Hordeum vulgare

Metals/Ions

Metals/Ions Comment Organism Structure
Mn2+ titration of periodate-oxidized oxalate oxidase with hydroxylamine completely eliminates the visible absorption, forming a homogeneous Mn2+ form of the enzyme. The fully reduced Mn2+ form lacks any detectable oxidase activity, reoxidation substantially restores the maximum activity. Hordeum vulgare
Mn3+ treatment of the periodate-oxidized enzyme with ascorbate results in a substantioal decrease in absorption, forming a complex that is spectroscopically identified as a Mn3+ species. Mn3+ form has a 5fold higher specific activity than native recombinant oxalate oxidase. Hordeum vulgare
Mn5+ titration of oxalate oxidase with sodium periodate results in nearly stoichometric oxidation of the enzyme to an intensely colored yellow complex, whose complete spectroscopic characterization lead to assignment to a superoxidized Mn5+ complex. Treatment of Mn2+ S49A oxalate oxidase generates the same yellow species as the glycosylated wild type enzyme. Mass spectra of isolated and periodate-treated oxalate oxidase are virtually identical, demonstating that no protein oxidation occurred. Peroxidate oxidation increases the specific activity about 5fold. Hordeum vulgare

Organism

Organism UniProt Comment Textmining
Hordeum vulgare
-
barley
-

Purification (Commentary)

Purification (Comment) Organism
-
Hordeum vulgare

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
the effect of the isotopic composition of the solvent is investigated by assaying oxalate oxidase in buffer prepared form H2O/D20 mixtures.The limiting values of Vs (steady state rate) lead to an estimate of the overall solvent kinetic isotope effect kH2O/kD2O = 8.5 (k=burst rate constant) Hordeum vulgare
additional information
-
Vimax (initial maximum velocity) is nearly independent of pH over the range from pH 3 to 5 Hordeum vulgare
21.9
-
native wild type oxalate oxidase Hordeum vulgare
94
-
native S49A oxalate oxidase Hordeum vulgare
139
-
periodate-oxidized oxalate oxidase, Mn5+ content 100% Hordeum vulgare
156
-
ascorbate-reduced oxalate oxidase, Mn3+ content above 95% Hordeum vulgare

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
9.7
-
oxalate oxalate oxidase activity is measured by oxygen uptake assay with a Clark oxygen electrode in a thermostated cell (25°C) Hordeum vulgare

pH Range

pH Minimum pH Maximum Comment Organism
2.5 6 the catalytic efficiency (Vmax/Km) increases continuously to lower pH Hordeum vulgare