Fig 3.

S. coelicolor ΔddlA is dependent on vancomycin for viability (A) but still produces cell wall PG precursors terminating in d-Ala-d-Ala (B). (A) The ΔddlA mutant strain requires the presence of vancomycin for growth, analogous to the ΔfemX mutant previously reported (8). The M600 parent strain is shown as a control. (B) HPLC-MS/MS analysis of cytoplasmic PG cell wall precursors isolated from the ΔddlA null mutant grown in the presence of 20 μg/ml vancomycin. The analysis was performed on biological replicates, and a representative result is shown. Peaks eluting after 11.92 min (peak 1) and 17.75 min (peak 2) using detection by UV absorbance at 254 nm were identified as UDP-MurNAc-pentapeptide (the d-Ala-d-Ala-containing PG precursor [1,193 Da]) and UDP-MurNAc-pentadepsipeptide (the d-Ala-d-Lac-containing PG precursor [1,194 Da]), respectively. uAU, microabsorbance units.