Title | Production, biophysical characterization and crystallization of Pseudomonas putida GraA and its complexes with GraT and the graTA operator. |
Publication Type | Journal Article |
Year of Publication | 2017 |
Authors | Talavera, A., H. Tamman, A. Ainelo, S. Hadǽi, A. Garcia-Pino, R. Hõrak, A. Konijnenberg, and R. Loris |
Journal | Acta Crystallogr F Struct Biol Commun |
Volume | 73 |
Issue | Pt 8 |
Pagination | 455-462 |
Date Published | 2017 Aug 01 |
ISSN | 2053-230X |
Keywords | Amino Acid Sequence, Antitoxins, Bacterial Proteins, Bacterial Toxins, Cloning, Molecular, Crystallization, Crystallography, X-Ray, Escherichia coli, Gene Expression, Gene Expression Regulation, Bacterial, Genetic Vectors, Operon, Protein Binding, Protein Multimerization, Pseudomonas putida, Recombinant Fusion Proteins, X-Ray Diffraction |
Abstract | The graTA operon from Pseudomonas putida encodes a toxin-antitoxin module with an unusually moderate toxin. Here, the production, SAXS analysis and crystallization of the antitoxin GraA, the GraTA complex and the complex of GraA with a 33 bp operator fragment are reported. GraA forms a homodimer in solution and crystallizes in space group P2, with unit-cell parameters a = 66.9, b = 48.9, c = 62.7 Å, β = 92.6°. The crystals are likely to contain two GraA dimers in the asymmetric unit and diffract to 1.9 Å resolution. The GraTA complex forms a heterotetramer in solution. Crystals of the GraTA complex diffracted to 2.2 Å resolution and are most likely to contain a single heterotetrameric GraTA complex in the asymmetric unit. They belong to space group P4 or P4, with unit-cell parameters a = b = 56.0, c = 128.2 Å. The GraA-operator complex consists of a 33 bp operator region that binds two GraA dimers. It crystallizes in space group P3 or P3, with unit-cell parameters a = b = 105.6, c = 149.9 Å. These crystals diffract to 3.8 Å resolution. |
DOI | 10.1107/S2053230X17009438 |
Alternate Journal | Acta Crystallogr F Struct Biol Commun |
PubMed ID | 28777088 |
Grant List | G.0135.15N / / Fonds Wetenschappelijk Onderzoek / G0C1213N / / Fonds Wetenschappelijk Onderzoek / UABR/11/012 / / The Hercules Foundation / IUT20-19 / / Estonian Research Council / PUT1351 / / Estonian Research Council / OZR2232 / / Vrije Universiteit Brussel / SPR13 / / Vrije Universiteit Brussel / |
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