Interactions in solution and crystallization of Aspergillus flavus urate oxidase

Francoise Bonnete, Denis Vivares, Christelle Robert, N. Colloc'h

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Interparticle interactions of urate oxidase from Aspergillus flavus have been studied by small-angle X-ray scattering to determine crystallization conditions. This enzyme is a homotetramer with a total molecular weight of 128 kDa. It is a slightly basic protein (pI between 7.5 and 8). The interaction potentials have been studied as a function of the main thermodynamic and chemical parameters: temperature, protein concentration, pH, salt nature and concentration, addition of polyols. In 10 mM sodium carbonate at pH 10.5, the interactions are slightly repulsive and become less repulsive with a pH closer to pI. With the addition of carbonate, the protein loses its tetrameric structure for a dimeric one; with formate, the tetrameric structure remains stable. We also studied the effect of polyethylene glycols as it had been done with high molecular weight proteins. With the addition of PEG 8 K, the interactions became less repulsive and even turned attractive with the addition of both PEG 8 K and salt. Protein crystals of urate oxidase were observed in slightly repulsive conditions (second virial coefficient A2 about +10−5 mol ml g−2 instead of –2 to –8×10−4 mol ml g−2 for low molecular weight proteins).
Original languageEnglish
Publication statusPublished - 15 Aug 2001


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