Crystallographic study of glutathione-dependent dehydroascorbate reductase from spinach chloroplasts

Abstract

Glutathione-dependent dehydroascorbate reductase (GSH-DHAR) catalyzes the reduction of dehydroascorbate to ascorbate using reduced glutathione as the electron donor. GSH-DHAR from spinach chloroplasts is considered to play a pivotal role to regenerate ascorbate that was oxidized to scavenge active oxygen species generated in the process of photosynthesis. Recombinant GSH-DHAR produced in Esherichia coli was crystallized by the hanging-drop vapour-diffusion method. The crystals were monoclinic, space group C2, with unit-cell dimensions a = 98.25, b = 39.96, c = 106.86 Å, ß = 110.46°. The asymmetric unit contains two molecules, giving a crystal volume per enzyme mass (Vm) of 2.06 Å3 Da-1 and a solvent content of 40.3%. A full set of X-ray diffraction data were collected to 2.2 Å Bragg spacing from three native crystals with an overall Rmerge of 6.5% and completeness of 93.4%.