Glycan strand cleavage by a lytic transglycosylase, MltD contributes to the expansion of peptidoglycan in Escherichia coli

Peptidoglycan (PG) is a protective sac-like exoskeleton present in most bacterial cell walls. It is a large, covalently cross-linked mesh-like polymer made up of several glycan strands cross-bridged to each other by short peptide chains. Because PG forms a continuous mesh around the bacterial cytoplasmic membrane, opening the mesh is critical to generate space for the incorporation of new material during its expansion. In Escherichia coli , the ‘space-making activity’ is known to be achieved by cleavage of cross-links between the glycan strands by a set of redundant PG endopeptidases whose absence leads to rapid lysis and cell death. Here, we demonstrate a hitherto unknown role of glycan strand cleavage in making space for cell wall expansion in E. coli . We find that overexpression of a membrane-bound lytic transglycosylase, MltD that cuts the glycan polymers of the PG sacculus rescues the cell lysis caused by the absence of essential cross-link specific endopeptidases, MepS, MepM and MepH. Further detailed genetic and biochemical analysis revealed that MltD works in conjunction with cross-link specific endopeptidases to expand the PG sacculus. Interestingly, we find that cellular MltD levels are stringently controlled by two independent regulatory pathways. MltD undergoes regulated proteolysis by NlpI-Prc, a periplasmic adaptor-protease complex that specifically degrades two of the elongation-specific endopeptidases, MepS and MepH. In addition, MltD levels are post-transcriptionally controlled by RpoS, a stationary-phase specific sigma factor. Overall, our results show that coordinated cleavage of the glycan strands and the peptide cross-bridges facilitates the opening of the PG mesh for successful expansion of the cell wall during growth of a bacterium.