Crystallization of Pseudomonas aeruginosa AmrZ protein: Development of a comprehensive method for obtaining and optimization of protein-DNA crystals

Abstract

The AmrZ protein from the pathogenic bacterium Pseudomonas aeruginosa is a transcription factor that activates and represses the genes for several potent virulence factors, which gives the bacteria a selective advantage in infection. AmrZ was crystallized in complex with DNA containing the amrZ1 repressor binding site. Obtaining crystals of the complex required the integration of a number of well known techniques along with the development of new methods. Here, these processes are organized and combined into a comprehensive method which yielded diffraction-quality crystals. Part of this method included thorough data mining of the crystallization conditions of protein-DNA complexes to create a new directed crystallization screen. An optimized technique for the verification of protein-DNA complexes in crystals is also presented. Taken together, the methods described in this article attempt to streamline the difficult process of obtaining diffraction-quality crystals of protein-DNA complexes through the organization of older methods combined with the introduction of new techniques. © 2012 International Union of Crystallography. All rights reserved.