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Literature summary for 4.1.1.2 extracted from

  • Saylor, B.T.; Reinhardt, L.A.; Lu, Z.; Shukla, M.S.; Nguyen, L.; Cleland, W.W.; Angerhofer, A.; Allen, K.N.; Richards, N.G.
    A structural element that facilitates proton-coupled electron transfer in oxalate decarboxylase (2012), Biochemistry, 51, 2911-2920.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 4.1.1.2

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of C-terminally His6-tagged wild-type enzyme and mutants T165V and T165S, expression is induced in the presence of 5 mM MnCl2 after heat shocking the bacteria for 18 min at 42°C Bacillus subtilis

Crystallization (Commentary)

Crystallization (Comment) Organism
purified His6-tagged enzyme mutant T165V, sitting drop vapor diffusion method, mixing of 0.0012 ml of 6 mg/mL protein in 1 mM HEPES buffer, pH 7.5, with 0.0012 ml of precipitant solution containing 10% w/v PEG 6000 and 2 M NaCl, and equilibration against well solution, 17°C, 1 week, X-ray diffraction structure determination and analysis at 2.31 A resolution Bacillus subtilis

Protein Variants

Protein Variants Comment Organism
T165S site-directed mutagenesis, the T165S enzyme variant exhibits similar catalytic activity to wild-type enzyme when adjusted for the amount of Mn(II) incorporated into the recombinant protein Bacillus subtilis
T165V site-directed mutagenesis, removal a conserved Arg/Thr hydrogen bonding interaction, the mutant exhibits impaired catalytic activity. The T165V OxDC variant exhibits a 15fold reduced catalytic efficiency and a lower level of oxalate consumption per dioxygen molecule compared to the wild-type Bacillus subtilis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information steady-state kinetics of the T165S and T165V enzyme mutants, overview Bacillus subtilis

Metals/Ions

Metals/Ions Comment Organism Structure
Mn2+ required, the conformational properties of an active-site loop segment, defined by residues Ser161-Glu162-Asn163-Ser164, are important for modulating the intrinsic reactivity of Mn(II) in the active site Bacillus subtilis

Organism

Organism UniProt Comment Textmining
Bacillus subtilis O34714
-
-
Bacillus subtilis 168 O34714
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant C-terminally His6-tagged enzyme mutants T165V and T165S by nickel affinity chromatography Bacillus subtilis

Reaction

Reaction Comment Organism Reaction ID
oxalate = formate + CO2 minimal, two step kinetic mechanism for enzyme-catalyzed decarboxylation proton-coupled electron transfer and decarboxylation Bacillus subtilis
a

Synonyms

Synonyms Comment Organism
OXDC
-
 Bacillus subtilis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
22
-
 assay at Bacillus subtilis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
4.2 5.7 assay at Bacillus subtilis

Cofactor

Cofactor Comment Organism Structure
O2 O2 might participate as a reversible electron sink in two putative proton-coupled electron transfer steps and is not merely used to generate a protein-based radical or oxidized metal center Bacillus subtilis

pI Value

Organism Comment pI Value Maximum pI Value
Bacillus subtilis recombinant enzyme mutant T165V has two isoelectric points at pH 4.2 and pH 5.7
-
 4.2
Bacillus subtilis recombinant enzyme mutant T165V has two isoelectric points at pH 4.2 and pH 5.7
-
 5.7

General Information

General Information Comment Organism
additional information the conformational properties of an active-site loop segment, defined by residues Ser161-Glu162-Asn163-Ser164, are important for modulating the intrinsic reactivity of Mn(II) in the active site of Bacillus subtilis oxalate decarboxylase. O2 might participate as a reversible electron sink in two putative proton-coupled electron transfer steps and is not merely used to generate a protein-based radical or oxidized metal center. The conserved Arg/Thr hydrogen bond is important for correctly locating the side chain of Glu162, which mediates a proton-coupled electron transfer step prior to decarboxylation in the catalytically competent form of the enzyme Bacillus subtilis