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Title: Crystallization and preliminary X-ray analysis of the complex between a Bacillus subtilis α/β-type small acid-soluble spore protein and DNA

Abstract

An α/β-type small, acid-soluble spore protein (SASP) from Bacillus subtilis, a major source of DNA protection against damaging effects in spores, was crystallized in a functionally relevant complex with a double-stranded DNA. This report provides insights into initial characterization of the complex and its structure elucidation. An engineered variant of an α/β-type small acid-soluble spore protein (SASP) from Bacillus subtilis was crystallized in a complex with a ten-base-pair double-stranded DNA by the hanging-drop vapor-diffusion method using ammonium sulfate as a precipitating agent. Crystals grew at 281 K using sodium cacodylate buffer pH 5.5 and these crystals diffracted X-rays to beyond 2.4 Å resolution using synchrotron radiation. The crystallized complex contains two or three SASP molecules bound to one DNA molecule. The crystals belong to the hexagonal space group P6{sub 1}22 or P6{sub 5}22, with unit-cell parameters a = b = 87.0, c = 145.4 Å, α = β = 90.0, γ = 120.0°. Diffraction data were 96.6% complete to 2.4 Å resolution, with an R{sub sym} of 8.5%. Structure solution by the multiwavelength/single-wavelength anomalous dispersion method using isomorphous crystals of selenomethionine-labeled protein is in progress.

Authors:
 [1]; ;  [2];  [3];  [1]
+ Show Author Affiliations
  1. Children’s Hospital Oakland Research Institute, Oakland, CA 94609 (United States)
  2. Department of Molecular, Microbial and Structural Biology, University of Connecticut Health Center, Farmington, CT 06032 (United States)
  3. Berkeley Center for Structural Biology, Lawrence Berkeley National Laboratory, Berkeley, CA 94720 (United States)
Publication Date:
OSTI Identifier:
22360347
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Crystallographica. Section F; Journal Volume: 63; Journal Issue: Pt 6; Other Information: PMCID: PMC2335083; PMID: 17554173; PUBLISHER-ID: ll5117; OAI: oai:pubmedcentral.nih.gov:2335083; Copyright (c) International Union of Crystallography 2007; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United Kingdom
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; AMMONIUM SULFATES; BUFFERS; CRYSTALLIZATION; CRYSTALS; DIFFRACTION; DIFFUSION; DNA; MOLECULES; RESOLUTION; SAFETY; SODIUM; SPACE GROUPS; SYNCHROTRON RADIATION; WAVELENGTHS

Citation Formats

Bumbaca, Daniela, Kosman, Jeffrey, Setlow, Peter, Henderson, R. Keith, and Jedrzejas, Mark J., E-mail: mjedrzejas@chori.org. Crystallization and preliminary X-ray analysis of the complex between a Bacillus subtilis α/β-type small acid-soluble spore protein and DNA. United Kingdom: N. p., 2007. Web. doi:10.1107/S1744309107022750.
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Bumbaca, Daniela, Kosman, Jeffrey, Setlow, Peter, Henderson, R. Keith, & Jedrzejas, Mark J., E-mail: mjedrzejas@chori.org. Crystallization and preliminary X-ray analysis of the complex between a Bacillus subtilis α/β-type small acid-soluble spore protein and DNA. United Kingdom. doi:10.1107/S1744309107022750.
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Bumbaca, Daniela, Kosman, Jeffrey, Setlow, Peter, Henderson, R. Keith, and Jedrzejas, Mark J., E-mail: mjedrzejas@chori.org. Fri . "Crystallization and preliminary X-ray analysis of the complex between a Bacillus subtilis α/β-type small acid-soluble spore protein and DNA". United Kingdom. doi:10.1107/S1744309107022750.
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@article{osti_22360347,
title = {Crystallization and preliminary X-ray analysis of the complex between a Bacillus subtilis α/β-type small acid-soluble spore protein and DNA},
author = {Bumbaca, Daniela and Kosman, Jeffrey and Setlow, Peter and Henderson, R. Keith and Jedrzejas, Mark J., E-mail: mjedrzejas@chori.org},
abstractNote = {An α/β-type small, acid-soluble spore protein (SASP) from Bacillus subtilis, a major source of DNA protection against damaging effects in spores, was crystallized in a functionally relevant complex with a double-stranded DNA. This report provides insights into initial characterization of the complex and its structure elucidation. An engineered variant of an α/β-type small acid-soluble spore protein (SASP) from Bacillus subtilis was crystallized in a complex with a ten-base-pair double-stranded DNA by the hanging-drop vapor-diffusion method using ammonium sulfate as a precipitating agent. Crystals grew at 281 K using sodium cacodylate buffer pH 5.5 and these crystals diffracted X-rays to beyond 2.4 Å resolution using synchrotron radiation. The crystallized complex contains two or three SASP molecules bound to one DNA molecule. The crystals belong to the hexagonal space group P6{sub 1}22 or P6{sub 5}22, with unit-cell parameters a = b = 87.0, c = 145.4 Å, α = β = 90.0, γ = 120.0°. Diffraction data were 96.6% complete to 2.4 Å resolution, with an R{sub sym} of 8.5%. Structure solution by the multiwavelength/single-wavelength anomalous dispersion method using isomorphous crystals of selenomethionine-labeled protein is in progress.},
doi = {10.1107/S1744309107022750},
journal = {Acta Crystallographica. Section F},
number = Pt 6,
volume = 63,
place = {United Kingdom},
year = {Fri Jun 01 00:00:00 EDT 2007},
month = {Fri Jun 01 00:00:00 EDT 2007}
}
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Journal Article:
DOI:  10.1107/S1744309107022750
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  • ; ; - Journal of Bacteriology; (USA)
    Small, acid-soluble proteins (SASP) of the alpha/beta-type are associated with DNA in spores of Bacillus subtilis. Induction of synthesis of alpha/beta-type SASP in Escherichia coli resulted in rapid cessation of DNA synthesis, followed by a halt in RNA and then protein accumulation, although significant mRNA and protein synthesis continued. There was a significant loss in viability associated with SASP synthesis in E. coli: recA+ cells became extremely long filaments, whereas recA mutant cells became less filamentous. The nucleoids of cells with alpha/beta-type SASP were extremely condensed, as viewed in both light and electron microscopes, and immunoelectron microscopy showed that themore » alpha/beta-type SASP were associated with the cell DNA. Induction of alpha/beta-type SASP synthesis in E. coli increased the negative superhelical density of plasmid DNA by approximately 20%; UV irradiation of E. coli with alpha/beta-type SASP gave reduced yields of thymine dimers but significant amounts of the spore photoproduct. These changes in E. coli DNA topology and photochemistry due to alpha/beta-type SASP are similar to the effects of alpha/beta-type SASP on the DNA in Bacillus spores, further suggesting that alpha/beta-type SASP are a major factor determining DNA properties in bacterial spores.« less

  • ; ; ; ... - Acta Crystallographica. Section F
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  • ; ; ; ... - Acta Crystallographica. Section F
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  • ; ; ; ... - Acta Crystallographica. Section F
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