Cloning, expression, purification, crystallization and preliminary X-ray diffraction studies of a major group 7 allergen, Der f 7, from the dust mite Dermatophagoides farinae

Abstract

Der f 7 is a major group 7 allergen from the dust mite Dermatophagoides farinae that shows 86% sequence identity to the homologous allergen Der p 7 from D. pteronyssinus. Der f 7 was successfully overexpressed in an Escherichia coli expression system and purified to homogeneity using Ni-NTA affinity and size-exclusion column chromatography. SeMet-labelled Der f 7 was crystallized by the hanging-drop vapour-diffusion method using a reservoir solution consisting of 0.1 M bis-tris pH 7.4 and 28% polyethylene glycol monomethyl ether 2000 at 293 K. X-ray diffraction data were collected to 2.24 Å resolution using synchrotron radiation. The crystals belonged to the orthorhombic system, space group P2(1)2(1)2(1), with unit-cell parameters a = 50.19, b = 58.67, c = 123.81 Å. Based on the estimated Matthews coefficient (2.16 Å(3) Da(-1)), two molecules of Der f 7 could be present in the asymmetric unit of the crystal lattice.