Protein expression: general procedure.

For expression of Staphylococcus and Streptococcus FtsW and PBPs, E. coli C43(DE3) containing the expression plasmid was grown in 1 L TB supplemented with kanamycin or carbenicillin at 37 ºC with shaking until OD₆₀₀ was 0.7–0.8. The culture was cooled to 20 ºC before inducing protein expression with 500 µM isopropyl β-D-1-thiogalactopyranoside (IPTG). Cells were harvested 18 h post-induction by centrifugation (4,200 x g, 15 min, 4 ºC) and the pellet was stored at −80 ºC.