Crystallization and preliminary X-ray diffraction studies of two thermostable α-galactosidases from glycoside hydrolase family 36

Abstract

-Galactosidases from thermophilic organisms have gained interest owing to their applications in the sugar industry. The -galactosidases AgaA, AgaB and AgaA A355E mutant from Geobacillus stearothermophilus have been overexpressed in Escherichia coli. Crystals of AgaB and AgaA A355E have been obtained by the vapour-diffusion method and synchrotron data have been collected to 2.0 and 2.8 Å resolution, respectively. Crystals of AgaB belong to space group I222 or I2 1 2 1 2 1 , with unit-cell parameters a = 87.5, b = 113.3, c = 161.6 Å . Crystals of AgaA A355E belong to space group P3 1 21 or P3 2 21, with unit-cell parameters a = b = 150.1, c = 233.2 Å .

The Molecular Mechanism of Thermostable α-Galactosidases AgaA and AgaB Explained by X-ray Crystallography and Mutational Studies

“…AgaA and AgaA A355E crystallize in the trigonal space group P3 2 21, and AgaB crystallizes in the orthorhombic space group I222 (21). The AgaA and AgaA A355E crystals contain a tetramer with 222-point group symmetry in their asymmetric unit.…”

“…Crystallization-AgaA A355E and AgaB were crystallized in different space groups as described previously (21). We tried to grow new AgaB crystals but were not successful.…”

The melREDCA Operon Encodes a Utilization System for the Raffinose Family of Oligosaccharides in Bacillus subtilis

“…The rate of melibiose hydrolysis was determined by assessing the amount of released glucose, which was determined by a glucose-hexokinase test (DiaSys GmbH, Holzheim, Germany) and quantified according to a glucose standard curve. The rates of raffinose and stachyose hydrolysis were measured by assessing the release of galactose after separation of the reaction mixture by high-performance liquid chromatography (HPLC), as described before (56). The hydrolysis of the mentioned sugars was measured from a final concentration of 100 mM under standard assay conditions.…”