FIG. 2.
Isolation of recombinant MreC/MreD and PBP2b/RodA complexes from pneumococcus. (A) MreC and MreD were expressed in Escherichia coli from an artificial operon fused with N-terminal Strep- and His8-tags, respectively. Membranes were isolated and solubilized with n-dodecyl-β-d-maltopyranoside. The complex was isolated by two successive affinity chromatography steps on Strep-Tactin® and Ni-NTA. (B) PBP2b with a C-terminal Strep-tag and RodA with an N-terminal His8-tag were similarly expressed and the complex isolated, in that case, first by Ni-NTA chromatography followed by a Strep-Tactin® purification. L, W, and E stand for load, wash, and elution. Samples were analyzed by Coomassie-stained sodium dodecyl sulfate–polyacrylamide gel electrophoresis.