Purification, crystallization and preliminary X-ray diffraction analysis of NADP-dependent glutamate dehydrogenase from Aspergillus niger

Glutamate dehydrogenase (GDH) catalyzes the NAD-dependent or NADP-dependent oxidative deamination of L-glutamate to 2-oxoglutarate and ammonia. This important reversible reaction establishes the link between carbon and nitrogen metabolism. In this study, Aspergillus niger NADP-GDH (AnGDH) has been overexpressed and purified. Purified AnGDH, with a high specific activity of 631.1 units per milligram of protein, was crystallized and the crystal diffracted to 2.9 Å resolution using a home X-ray source. Preliminary analysis of the X-ray diffraction data showed that the crystal belonged to space group R32, with unit-cell parameters a = b = 173.8, c = 241.5 Å, α = β = 90, γ = 120°. The crystals exhibited an unusually high solvent content (83.0%) and had only one molecule in the asymmetric unit. Initial phases were obtained by molecular replacement, and model building and structure refinement of AnGDH are in progress.