Figure 1. PGRPs inhibit intracellular step in peptidoglycan synthesis and localize to the newly formed cell separation site.

(a) PGRPs inhibit peptidoglycan synthesis in S. aureus measured by incorporation of ³H-GlcNAc into insoluble peptidoglycan; 0.5 M sucrose prevents osmotic lysis; means ± SEM of 3 – 4 experiments. (b) PGRPs do not inhibit transpeptidation in S. aureus measured by incorporation of ³H-GlcNAc into secreted uncrosslinked polymeric peptidoglycan. (c) PGRPs do not inhibit transglycosylation in S. aureus measured as accumulation of ³H-GlcNAc-lipid II in cell membrane; means ± SEM of 3–4 (b) or 4–5 (c) experiments; *, P<0.03; **, P<0.0002 (t-test). (d) PGRPs localize to the newly formed cell separation site and not to the sites of peptidoglycan synthesis in B. subtilis (top rows) or S. aureus (bottom rows); different stages of cell division shown; each panel from left to right shows green (BODIPY-FL-vancomycin), red (Alexa Fluor 594-PGLYRP-4), overlay red and green, and overlay of red and dark field view. (e) PGLYRP-1 or PGLYRP-4 (red) co-localize with LytE and LytF (green) in B. subtilis and LytE/LytF peptide inhibits detection LytE and LytF; each panel from left to right shows red (PGRP), green (LytE and LytF), overlay red and green (yellow), and overlay of red and green and dark field view; bar = 1 μm. (f) LytE- and LytF-generated cell separation sites are required for efficient killing of B. subtilis by PGRPs, shown by lower killing of ΔlytE:ΔlytF or ΔlytE: ΔlytF: ΔcwlS mutants than of WT bacteria by PGLYRP-1; means ± SEM of 3 experiments; *, P<0.05 versus WT (t-test).