Purification, crystallization and preliminary X‐ray diffraction studies of the ATP‐binding subunit of an ABC transporter from Geobacillus kaustophilus

ATP‐binding cassette (ABC) transporters, also known as traffic ATPases, form a large family of integral membrane proteins responsible for the translocation of a variety of chemically diverse substrates across the lipid bilayers of cellular membranes of both prokaryotes and eukaryotes by the hydrolysis of ATP. The ATP‐binding subunit of an ABC transporter from Geobacillus kaustophilus , a homodimeric enzyme, was overexpressed in Escherichia coli and purified. Crystals were obtained using the microbatch‐under‐oil method at 291 K. X‐ray diffraction data to 1.6 Å resolution were collected on SPring‐8 beamline BL26B1. The crystals belonged to the orthorhombic space group I 222, with unit‐cell parameters a = 54.94, b  = 78.63, c = 112.96 Å. Assuming the presence of a dimer in the asymmetric unit gave a crystal volume per protein weight ( V M ) of 2.32 Å 3  Da −1 and a solvent content of 47%; this was consistent with the results of a dynamic light‐scattering experiment, which showed a dimeric state of the protein in solution. Molecular‐replacement trials using the crystal structure of HisP from the Salmonella typhimurium ATP‐binding subunit of an ABC transporter as a search model did not provide a satisfactory solution, indicating that the two ATP‐binding subunits of ABC transporters have substantially different structures.