A1 Refereed original research article in a scientific journal
Overexpression, purification, crystallization and preliminary X-ray diffraction analysis of the F1 antigen Caf1M-Caf1 chaperone-subunit pre-assembly complex from Yersinia pestis
Authors: Zavialov A, Berglund J, Knight SD
Publisher: INT UNION CRYSTALLOGRAPHY
Publication year: 2003
Journal: Acta crystallographica section D : structural biology
Journal name in source: ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY
Journal acronym: ACTA CRYSTALLOGR D
Volume: 59
First page : 359
Last page: 362
Number of pages: 4
ISSN: 2059-7983
DOI: https://doi.org/10.1107/S0907444902021054
Abstract
The F1 capsular antigen of the plague-causing pathogen Yersinia pestis is assembled from monomeric Caf1 subunits via the Caf1M/Caf1A chaperone/usher system. Y. pestis Caf1M-Caf1 chaperone-subunit complex was purified from the periplasm of Escherichia coli cells overexpressing Caf1M and Caf1 and was crystallized in PEG 4000 solution using hanging-drop vapour diffusion. The crystals diffract to a minimum Bragg spacing of 1.8 Angstrom and belong to space group P2(1), with unit-cell parameters a = 36.0, b = 69.2, c = 69.1 Angstrom, beta = 93.0degrees. SeMet-labelled Caf1M-Caf1 complexes were purified and crystallized under the same conditions. The SeMet crystals were identical to the native crystals and diffracted to 1.9 Angstrom. Heavy-atom derivative crystals were prepared by soaking in 10 mM K2PtCl4, giving two Pt sites per complex. The experimental electron-density map was obtained by a combination of MAD and MIR methods using both Se- and Pt-derivative crystals.
The F1 capsular antigen of the plague-causing pathogen Yersinia pestis is assembled from monomeric Caf1 subunits via the Caf1M/Caf1A chaperone/usher system. Y. pestis Caf1M-Caf1 chaperone-subunit complex was purified from the periplasm of Escherichia coli cells overexpressing Caf1M and Caf1 and was crystallized in PEG 4000 solution using hanging-drop vapour diffusion. The crystals diffract to a minimum Bragg spacing of 1.8 Angstrom and belong to space group P2(1), with unit-cell parameters a = 36.0, b = 69.2, c = 69.1 Angstrom, beta = 93.0degrees. SeMet-labelled Caf1M-Caf1 complexes were purified and crystallized under the same conditions. The SeMet crystals were identical to the native crystals and diffracted to 1.9 Angstrom. Heavy-atom derivative crystals were prepared by soaking in 10 mM K2PtCl4, giving two Pt sites per complex. The experimental electron-density map was obtained by a combination of MAD and MIR methods using both Se- and Pt-derivative crystals.
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