Isolation and characterisation of EfeM, a periplasmic component of the putative EfeUOBM iron transporter of Pseudomonas syringae pv. syringae.
The EfeM protein is a component of the putative EfeUOBM iron-transporter of Pseudomonas syringae pathovar syringae and is thought to act as a periplasmic, ferrous-iron binding protein. It contains a signal peptide of 34 amino acid residues and a C-terminal 'Peptidase_M75' domain of 251 residues. The C-terminal domain contains a highly conserved 'HXXE' motif thought to act as part of a divalent cation-binding site. In this work, the gene (efeM or 'Psyr_3370') encoding EfeM was cloned and over-expressed in Escherichia coli, and the mature protein was purified from the periplasm. Mass spectrometry confirmed the identity of the protein (M(W) 27,772Da). Circular dichroism spectroscopy of EfeM indicated a mainly alpha-helical structure, consistent with bioinformatic predictions. Purified EfeM was crystallised by hanging-drop vapor diffusion to give needle-shaped crystals that diffracted to a resolution of 1.6A. This is the first molecular study of a peptidase M75 domain with a presumed iron transport role.
Rajasekaran
Mohan B
MB
School of Biological Sciences Harborne Building, Whiteknights Campus, Reading, RG6 6AS, UK.
Mitchell
Sue A
SA
Gibson
Trevor M
TM
Hussain
Rohanah
R
Siligardi
Giuliano
G
Andrews
Simon C
SC
Watson
Kimberly A
KA
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D027682:Cation Transport Proteins / Q000737:chemistry* / Q000235:genetics; D018360:Crystallography, X-Ray; D017136:Ion Transport; D007501:Iron / Q000378:metabolism*; D033903:Periplasmic Proteins / Q000737:chemistry* / Q000235:genetics; D017434:Protein Structure, Tertiary; D044224:Pseudomonas syringae / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization
D001426:Bacterial Proteins; D027682:Cation Transport Proteins; D033903:Periplasmic Proteins; D007501:Iron