Expression, purification, crystallization and preliminary X-ray diffraction analysis of the novel modular DNA-binding protein BurrH in its apo form and in complex with its target DNA

Stefano Stella; Rafael Molina; Claudia Bertonatti; Alexandre Juillerrat; Guillermo Montoya

doi:10.1107/s2053230x13033037

Expression, purification, crystallization and preliminary X-ray diffraction analysis of the novel modular DNA-binding protein BurrH in its apo form and in complex with its target DNA

Stefano Stella, Rafael Molina, Claudia Bertonatti, Alexandre Juillerrat, Guillermo Montoya^*

^*Corresponding author af dette arbejde

Protein Structure and Function Program

8 Citationer (Scopus)

Abstract

Different genome-editing strategies have fuelled the development of new DNA-targeting molecular tools allowing precise gene modifications. Here, the expression, purification, crystallization and preliminary X-ray diffraction of BurrH, a novel DNA-binding protein from Burkholderia rhizoxinica, are reported. Crystallization experiments of BurrH in its apo form and in complex with its target DNA yielded crystals suitable for X-ray diffraction analysis. The crystals of the apo form belonged to the primitive hexagonal space group P31 or its enantiomorph P32, with unit-cell parameters a = b = 73.28, c = 268.02 Å, α = β = 90, γ = 120°. The BurrH-DNA complex crystallized in the monoclinic space group P2₁, with unit-cell parameters a = 70.15, b = 95.83, c = 76.41 Å, α = γ = 90, β = 109.51°. The self-rotation function and the Matthews coefficient suggested the presence of two protein molecules per asymmetric unit in the apo crystals and one protein-DNA complex in the monoclinic crystals. The crystals diffracted to resolution limits of 2.21 and 2.65 Å, respectively, using synchrotron radiation.

Originalsprog	Engelsk
Tidsskrift	Acta Crystallographica Section F:Structural Biology Communications
Vol/bind	70
Udgave nummer	1
Sider (fra-til)	87-91
Antal sider	5
ISSN	2053-230X
DOI	https://doi.org/10.1107/s2053230x13033037
Status	Udgivet - 1 jan. 2014

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10.1107/s2053230x13033037

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Stella, S., Molina, R., Bertonatti, C., Juillerrat, A., & Montoya, G. (2014). Expression, purification, crystallization and preliminary X-ray diffraction analysis of the novel modular DNA-binding protein BurrH in its apo form and in complex with its target DNA. Acta Crystallographica Section F:Structural Biology Communications, 70(1), 87-91. https://doi.org/10.1107/s2053230x13033037

Expression, purification, crystallization and preliminary X-ray diffraction analysis of the novel modular DNA-binding protein BurrH in its apo form and in complex with its target DNA. / Stella, Stefano ; Molina, Rafael; Bertonatti, Claudia et al.

I: Acta Crystallographica Section F:Structural Biology Communications, Bind 70, Nr. 1, 01.01.2014, s. 87-91.

Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review

Stella, S , Molina, R, Bertonatti, C, Juillerrat, A & Montoya, G 2014, 'Expression, purification, crystallization and preliminary X-ray diffraction analysis of the novel modular DNA-binding protein BurrH in its apo form and in complex with its target DNA', Acta Crystallographica Section F:Structural Biology Communications, bind 70, nr. 1, s. 87-91. https://doi.org/10.1107/s2053230x13033037

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abstract = "Different genome-editing strategies have fuelled the development of new DNA-targeting molecular tools allowing precise gene modifications. Here, the expression, purification, crystallization and preliminary X-ray diffraction of BurrH, a novel DNA-binding protein from Burkholderia rhizoxinica, are reported. Crystallization experiments of BurrH in its apo form and in complex with its target DNA yielded crystals suitable for X-ray diffraction analysis. The crystals of the apo form belonged to the primitive hexagonal space group P31 or its enantiomorph P32, with unit-cell parameters a = b = 73.28, c = 268.02 {\AA}, α = β = 90, γ = 120°. The BurrH-DNA complex crystallized in the monoclinic space group P21, with unit-cell parameters a = 70.15, b = 95.83, c = 76.41 {\AA}, α = γ = 90, β = 109.51°. The self-rotation function and the Matthews coefficient suggested the presence of two protein molecules per asymmetric unit in the apo crystals and one protein-DNA complex in the monoclinic crystals. The crystals diffracted to resolution limits of 2.21 and 2.65 {\AA}, respectively, using synchrotron radiation.",

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Expression, purification, crystallization and preliminary X-ray diffraction analysis of the novel modular DNA-binding protein BurrH in its apo form and in complex with its target DNA

Abstract

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