Figure 2.

Phenotypic characterization of strains overexpressing fmhA and fmhC. A, overnight cultures of WT strain Newman carrying control vector pitet, pitet-fmhA, or pitet-fmhC were treated with lysostaphin, and lysis was recorded as a decline in A₆₀₀ over time as described for Fig. 1C. B, peptidoglycan prepared from the same strains grown to A₆₀₀ 0.8–1.0 was treated with mutanolysin, and products were resolved by HPLC as described for Fig. 1D. Peaks denoted with Roman numerals (i–iv) were subjected to MALDI-TOF analysis and m/z values of the corresponding species are listed in Table 1. Shaded area under curves between 110–130 min highlights higher order cross-linking of peptidoglycan fragments (47). C, MALDI-TOF spectra of species eluting in peak i shown in panel C. D, Edman degradation profile of the major peptidoglycan fragment in peak i of the samples overexpressing fmhA and fmhC. The yield of released phenylthiohydantoin (PT)-modified amino acids is shown for five cleavage cycles.