Purification and characterization of recombinant EssB and truncated variants. (A) Diagrammatic representation of full length EssB and truncated variants produced in E. coli. Numbers indicate amino acid positions in the primary sequence and the grey box labeled PTMD depicts the hydrophobic sequence. (B) Coomassie gel of purified recombinant proteins as shown in panel A. Proteins were purified from E. coli by affinity chromatography and affinity tags were removed. (C) Size exclusion chromatography of full length EssB and truncated variants shown in panel B. Proteins (~100 μg) were loaded onto a SuperdexTM 75 10/300 GL and fractions (0.5 ml) were collected and analyzed by SDS-PAGE. Proteins in the gel were visualized by Coomassie staining. Masses of protein standards used for calibration are shown above the gels (158, 75, 43, 17 kDa) and correspond to the exclusion volumes of Aldolase, Conalbumin, Ovalbumin and Myoglobin, respectively. (D-E) TEM of purified recombinant EssB (D) and EssBΔM (E). The proteins were allowed to bind to glow discharged grids and were negatively stained using 2% uranyl acetate. This analysis reveals a rod-like structure for EssB and more spherical, aggregated-like structure for EssBΔM. Scale bar = 20 nm.