Crystal engineering: deletion mutagenesis of the 24 kDa fragment of the DNA gyrase B subunit from Staphylococcus aureus

• Published in: Acta crystallographica. Section D, Biological crystallography. Biological crystallography., 1999-09, Vol.55 (9), p.1626-1629
• Main Authors: Dale, Glenn E., Kostrewa, Dirk, Gsell, Bernard, Stieger, Martin, D'Arcy, Allan
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - biosynthesis; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - isolation & purification; crystal engineering; Crystallization; Crystallography, X-Ray; DNA Gyrase; DNA Topoisomerases, Type II - biosynthesis; DNA Topoisomerases, Type II - chemistry; DNA Topoisomerases, Type II - genetics; DNA Topoisomerases, Type II - isolation & purification; Escherichia coli - enzymology; Escherichia coli - genetics; Gene Deletion; Mutagenesis; Peptide Fragments - biosynthesis; Peptide Fragments - chemistry; Peptide Fragments - genetics; Peptide Fragments - isolation & purification; Staphylococcus aureus - enzymology
• ISSN: 1399-0047; 0907-4449; 1399-0047
• DOI: 10.1107/S0907444999008227

The 24 kDa fragment of DNA gyrase B from Staphylococcus aureus was expressed in Escherichia coli and purified for crystallization. Crystals of the wild‐type protein grew in the presence of cyclothialidine but proved difficult to reproduce. In order to improve the crystallization, the flexible regions of the protein were deleted by mutagenesis. The mutant proteins were analyzed by differential scanning calorimetry and the most stable mutants produced crystals. It was possible to reproducibly grow in the microbatch system single well defined crystals which belonged to the space group C2 and diffracted isotropically to approximately 2 Å resolution.