Expression purification crystallization and preliminary X-ray analysis of Pseudomonas aeruginosa PelD

摘要

The production of the PEL polysaccharide in Pseudomonas aeruginosa requires the binding of bis-(3′,5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) to the cytoplasmic GGDEF domain of the inner membrane protein PelD. Here, the overexpression, purification and crystallization of a soluble construct of PelD that encompasses the GGDEF domain and a predicted GAF domain is reported. Diffraction-quality crystals were grown using the hanging-drop vapour-diffusion method. The crystals grew as flat plates, with unit-cell parameters a = 88.3, b = 114.0, c = 61.9 Å, α = β = γ = 90.0°. The PelD crystals exhibited the symmetry of space group P21212 and diffracted to a minimum d-spacing of 2.2 Å. On the basis of the Matthews coefficient (V M = 2.29 ų Da⁻¹), it was estimated that two molecules are present in the asymmetric unit.