Expression, purification and crystallization of two peroxisomal acyl-CoA oxidases from Arabidopsis thaliana

Authors: Pedersen, Lise; Henriksen, Anette

Source: Acta Crystallographica Section D, Volume 60, Number 6, June 2004 , pp. 1125-1128(4)

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Abstract:

Two members of the acyl-CoA oxidase family from Arabidopsis thaliana have been cloned, overexpressed, purified and crystallized. Long-chain-specific acyl-CoA oxidase 1 crystals are characterized by a large variation in diffraction quality and non-isomorphous unit-cell parameters. The best crystals diffract to 2.0 Å using synchrotron radiation, have unit-cell parameters a = 85.2, b = 117.0, c = 131.0 Å, α = β = γ = 90° and show P2₁2₁2₁ symmetry. There are two polypeptide chains in the asymmetric unit. Short-chain-specific acyl-CoA oxidase 4 crystals are trigonal, space group P3₁21/P3₂21, with unit-cell parameters a = b = 198.7, c = 149.6 Å. The crystals are most likely to contain six polypeptide chains in the asymmetric unit. Freshly prepared acyl-CoA oxidase 4 crystals diffract to 3.9 Å at cryogenic temperature at beamline I711, Max-Lab, but the diffraction quality degenerates after storage for only a few days in the crystallization drop. A selenomethionine-substituted form of the protein was produced and two-wavelength MAD data were collected at beamline BW7A, EMBL Outstation, Hamburg.

Keywords: acyl-CoA oxidases

Document Type: Research article

DOI: http://dx.doi.org/10.1107/S0907444904007577

Publication date: 2004-06-01