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 Crystallogr F Struct Biol Commun. 2014 Jan;70(Pt 1):87-91. doi: 10.1107/S2053230X13033037. Epub 2013 Dec 24.

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 P3(1) or its enantiomorph P3(2), 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 P2(1), 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.

Keywords: Burkholderia rhizoxinica; BurrH; gene targeting; protein–DNA interaction.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoproteins / chemistry*
  • Apoproteins / isolation & purification
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / isolation & purification
  • Burkholderia / enzymology*
  • Crystallization
  • DNA / chemistry*
  • DNA / metabolism
  • DNA-Binding Proteins / chemistry*
  • DNA-Binding Proteins / isolation & purification*
  • X-Ray Diffraction*

Substances

  • Apoproteins
  • Bacterial Proteins
  • DNA-Binding Proteins
  • DNA