Oxalate Decarboxylase
Oxalate undergoes at least three types of metabolism in various organisms, all of which lead to decarboxylation but by very different mechanisms and with the formation of very different products. Oxalate decarboxylase (OxDC; EC 4.1.1.23) is found in fungi and bacteria. It catalyzes the decarboxylation to form carbon dioxide and formate according to eq. 8-7.
Oxalate oxidase (EC 1.2.3.4) catalyzes the reaction of oxalate with molecular oxygen to form two moles of carbon dioxide and a mole of hydrogen peroxide. Oxalyl CoA decar-boxylase (4.1.1.8) is a TPP-dependent enzyme that catalyzes the conversion of oxalyl CoA to carbon dioxide and formyl CoA.
OxDC is a manganese-enzyme that requires molecular oxygen to activate it but does not use it as a substrate. The main mechanistic information comes from heavy atom kinetic isotope effects and the crystal structure. The structure of the enzyme from Bacillus subtilis is shown in fig. 8-18 (Anand et al., 2002). The enzyme is a 264-kDa hexamer, in which each subunit is composed of two similar but not identical domains. In original structure shown in fig. 8-18, each domain incorporated a manganese binding site consisting of three histidine residues and a glutamate. Site b in the C-terminal domain included a second
Fig. 8-18. Bacillus subtilis oxalate decarboxylase is a homohexameric enzyme and is depicted in the top panel in stereo, with one subunit drawn in brown ribbons and the other five subunits drawn as gray Ca traces. The middle panel shows a single subunit with its bicupin fold (one cupin fold surrounds site a, and the other cupin fold surrounds site b). Three divalent metal ions are bound to each subunit in three distinct sites: a, b, and c. Manganese is bound to sites a and b and magnesium to site c. Formate was also observed bound to the metal ions in sites a and c. Each site is shown schematically in the bottom panel. Site b has been proposed to support oxalate decarboxylase activity because mutation of Glu333 results in a 25-fold activity drop (1.9-A resolution; PDB 1L3J; Anand et al., 2002).
Fig. 8-18. Bacillus subtilis oxalate decarboxylase is a homohexameric enzyme and is depicted in the top panel in stereo, with one subunit drawn in brown ribbons and the other five subunits drawn as gray Ca traces. The middle panel shows a single subunit with its bicupin fold (one cupin fold surrounds site a, and the other cupin fold surrounds site b). Three divalent metal ions are bound to each subunit in three distinct sites: a, b, and c. Manganese is bound to sites a and b and magnesium to site c. Formate was also observed bound to the metal ions in sites a and c. Each site is shown schematically in the bottom panel. Site b has been proposed to support oxalate decarboxylase activity because mutation of Glu333 results in a 25-fold activity drop (1.9-A resolution; PDB 1L3J; Anand et al., 2002).
glutamate residue in position to function as a catalyst (Glu333), and this site was thought to be the functional active site. Site a in the N-terminal domain included Leu153 in the position corresponding to Glu333 in the b site, and site a was thought not to be functional. In a later structure, sites a and b were found to be identical, with a catalytic glutamate in each site (Just et al., 2004). Magnesium is bound in a third site, c, that is unlike sites a and b.
The pH dependence for log V/K shows a single break downward with increasing pH, corresponding to a pKa of 4.2, exactly the value for the first ionization of oxalate, and leading to the conclusion that the substrate is hydrogen oxalate, HOOC-COO- (Reinhardt et al., 2003). The reaction proceeds without significant exchange of solvent oxygen into the unreacted oxalate or the formate produced.
HOV His.
Mn"
His | NGlu His
Glu333
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