Crystallization and preliminary X-ray diffraction analysis of the hyperthermostable NAD-dependent glutamate dehydrogenase from Pyrobaculum islandicum

NAD-dependent glutamate dehydrogenase from the hyperthermophilic archaeon Pyrobaculum islandicum was crystallized in the apo- and holoenzyme forms. Crystals were obtained using 2-propanol and polyethylene glycol MME 550 as precipitants for the apoenzyme and holoenzyme, respectively. The apoenzyme crystals belong to the trigonal space group P3(1)21 or its enantiomorph P3(2)21. The asymmetric unit contains three subunits; the values of the Matthews coef'cient (V-M) and the solvent content are 2.9 Angstrom(3) Da(-1) and 57%, respectively. A native data set was collected to a highest resolution limit of 4.0 Angstrom on an in-house X-ray source using a rotating-anode generator (overall R-sym of 12.3% and completeness of 97%). The holoenzyme crystals belong to the orthorhombic space group P2(1)2(1)2(1); the asymmetric unit contains one hexamer, giving a VM of 2.79 Angstrom(3) Da(-1) and a solvent content of 55%. Native and derivative data sets were collected. The crystals diffract to a maximum resolution of 2.8 Angstrom on the KEK-NW12 beamline at the Photon Factory and gave a data set with an overall R-sym of 7.9% and a completeness of 91%. Attempts are being made to solve the structure by the SIRAS method.