Purification, crystallization and preliminary X-ray diffraction studies of recombinant class A non-specific acid
R D Makde1, Vinay Kumar, A S Rao
1Synchrotron Radiation Section, Bhabha Atomic Research Centre, Mumbai 400 085, India.
Abstract:
The phoN gene of Salmonella enterica sv. Typhimurium strain MD6001 was cloned in the multicopy plasmid pBluescript SK(-). The nucleotide sequence of the cloned gene differs from the corresponding S. typhimurium LT2 sequence at 23 residues, leading to 15 amino-acid differences, but was very close to the S. typhi phoN sequence (only three nucleotide and two amino-acid differences). The recombinant PhoN protein was purified to homogeneity. Two forms of crystals were harvested from a single crystallization condition. Diffraction intensity data were collected using a laboratory X-ray source to resolution limits of 2.5 and 2.8 A for crystals belonging to space group C2 and C222(1), respectively. Based on non-crystallographic symmetry, four monomers of PhoN are expected to be present in the asymmetric unit of the C2 unit cell. Two monomers of a biologically active dimer in the asymmetric unit of the C222(1) unit cell are expected from the Matthews coefficient.
