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Oxygen-dependent coproporphyrinogen-III oxidase from Escherichia coli: one-step purification and biochemical
Sofia Macieira1, Berta M Martins, Robert Huber
1Max-Planck-Institut für Biochemie, Abt. Strukturforschung, Am Klopferspitz, 18-a, 82152, Martinsried, Germany. maciera@biochem.mpg.de
Abstract:
Coproporphyrinogen-III oxidase (CPO) catalyses the conversion of coproporphyrinogen-III to protoporphyrinogen-IX in the haem biosynthetic pathway, and its deficient activity is associated with human hereditary coproporphyria. The 47% sequence identity between the oxygen-dependent CPO from Escherichia coli and its human counterpart makes the bacterial enzyme a good model system for structural studies of this disease. Therefore, we overexpressed and purified to homogeneity the oxygen-dependent CPO from E. coli and its selenomethionine derivative fused with a His(6)-tag. Both preparations showed a specific activity of 37500 U mg(-1), had a subunit molecular mass of 35 kDa and behaved as a compact shaped dimer. First crystallisation trials produced plate-shaped diffracting crystals.