Crystallization and preliminary X-ray diffraction analysis of a dihaem cytochrome c peroxidase from Paracoccus denitrificans

Authors: Echalier, Aude; Goodhew, Celia F.; Pettigrew, Graham W.; Fülöp, Vilmos

Source: Acta Crystallographica Section D, Volume 60, Number 2, 1 February 2004 , pp. 331-333(3)

Publisher: Blackwell Publishing

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Abstract:

Cytochrome c peroxidase was isolated from Paracoccus denitrificans and purified to homogeneity in three steps prior to crystallization. Two different diffraction-quality crystal forms were obtained by the hanging-drop vapour-diffusion method using a number of screening conditions. The best (needle-shaped) crystal form is suitable for structural studies and was grown from solutions containing 20% PEG 8000, 0.1 M Tris pH 8.5 and 0.2 M MgCl₂. Crystals grew to a maximum length of approximately 0.7 mm and belong to the primitive monoclinic space group P2₁, with unit-cell parameters a = 78.3, b = 51.0, c = 167.2 Å, β = 97.9°. After a dehydration step and extensive optimization of the cryocooling conditions, a complete data set was collected to 2.2 Å from a native crystal of the fully oxidized form of the enzyme using synchrotron radiation.

Keywords: cytochrome c peroxidase; haem protein; calcium binding; crystal dehydration

Document Type: Research article

DOI: 10.1107/S0907444903026519

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