| Article ID | Journal | Published Year | Pages | File Type |
|---|---|---|---|---|
| 1931549 | Biochemical and Biophysical Research Communications | 2010 | 6 Pages |
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 (MW 27,772 Da). Circular dichroism spectroscopy of EfeM indicated a mainly α-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 1.6 Å. This is the first molecular study of a peptidase M75 domain with a presumed iron transport role.
Research highlights► Bioinformatic analysis reveals EfeM is a metallopeptidase with conserved HXXE motif. ► Mass spectrometry confirms EfeM consists of 251 residues, molecular weight 27,772Da. ► SRCD spectroscopy shows an α-helical secondary structure. ► Single crystals of EfeM are orthorhombic and diffract to 1.6Å resolution. ► Space group is P22121 with cell dimensions a = 46.74, b = 95.17 and c = 152.61 Å.
