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Acta Crystallogr F Struct Biol Commun. 2017 Aug 1;73(Pt 8):455-462. doi: 10.1107/S2053230X17009438. Epub 2017 Jul 26.

Production, biophysical characterization and crystallization of Pseudomonas putida GraA and its complexes with GraT and the graTA operator.

Author information

1
Structural Biology Brussels, Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussels, Belgium.
2
Institute of Molecular and Cell Biology, University of Tartu, 51010 Tartu, Estonia.
3
Institut de Biologie et de Médecine Moléculaires, Université Libre de Bruxelles, 6041 Gosselies, Belgium.

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 P21, 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 P41 or P43, 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 P31 or P32, with unit-cell parameters a = b = 105.6, c = 149.9 Å. These crystals diffract to 3.8 Å resolution.

KEYWORDS:

GraA; GraT; Pseudomonas putida; macromolecular complex; persistence; protein–DNA complex; ribosome biogenesis; toxin–antitoxin module

PMID:
28777088
PMCID:
PMC5544002
DOI:
10.1107/S2053230X17009438
[Indexed for MEDLINE]
Free PMC Article

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