The Redundancy of Peptidoglycan Carboxypeptidases Ensures Robust Cell Shape Maintenance in Escherichia coli

Abstract

Peptidoglycan (PG) is an essential structural component of the bacterial cell wall and maintains the integrity and shape of the cell by forming a continuous layer around the cytoplasmic membrane. The thin PG layer of Escherichia coli resides in the periplasm, a unique compartment whose composition and pH can vary depending on the local environment of the cell. Hence, the growth of the PG layer must be sufficiently robust to allow cell growth and division under different conditions. We have analyzed the PG composition of 28 mutants lacking multiple PG enzymes (penicillin-binding proteins [PBPs]) after growth in acidic or near-neutral-pH media. Statistical analysis of the muropeptide profiles identified dd-carboxypeptidases (DD-CPases) that were more active in cells grown at acidic pH. In particular, the absence of the DD-CPase PBP6b caused a significant increase in the pentapeptide content of PG as well as morphological defects when the cells were grown at acidic pH. Other DD-CPases (PBP4, PBP4b, PBP5, PBP6a, PBP7, and AmpH) and the PG synthase PBP1B made a smaller or null contribution to the pentapeptide-trimming activity at acidic pH. We solved the crystal structure of PBP6b and also demonstrated that the enzyme is more stable and has a lower Km at acidic pH, explaining why PBP6b is more active at low pH. Hence, PBP6b is a specialized DD-CPase that contributes to cell shape maintenance at low pH, and E. coli appears to utilize redundant DD-CPases for normal growth under different conditions.

Novel RpoS-Dependent Mechanisms Strengthen the Envelope Permeability Barrier during Stationary Phase

“…The different proteins with this activity are thought to act under different conditions. For instance, during active growth in neutral conditions, PBP5 is the main protein responsible for DD-CPase activity, while at a pH of 5, PBP6b is mainly responsible for DD-CPase activity (88). In addition, while PBP5 is most active in early log phase, PBP6 and PBP6b are thought to be most active in mid-log and stationary phases (83,89).…”

Peptidoglycan in obligate intracellular bacteria

“…Nascent or polymerised peptidoglycan is often subject to secondary modifications, such as N-deacetylation of residues in glycan chains, and trimming of the peptides by carboxypeptidases (Vollmer, 2008;Peters et al, 2016). Other peptidoglycan hydrolases cleave at various sites in the glycan and peptide components of peptidoglycan and these activities cause a release of peptidoglycan fragments from the sacculus (peptidoglycan turnover) (Vollmer et al, 2008b).…”

Building peptidoglycan inside eukaryotic cells: A view from symbiotic and pathogenic bacteria

“…An example is the d,d-carboxypeptidase PBP6b of E. coli. Unlike other d,d-carboxypeptidases like PBP4, PBP5, PBP6 and PBP7; PBP6b is the only capable of suppressing a cell shape defect at acidic pH (Peters et al, 2016).…”

“…An example is the d , d ‐carboxypeptidase PBP6b of E. coli . Unlike other d , d ‐carboxypeptidases like PBP4, PBP5, PBP6 and PBP7; PBP6b is the only capable of suppressing a cell shape defect at acidic pH (Peters et al, ). Another recent study involving complementation of an E. coli PBP5‐null mutant with other d , d ‐carboxypeptidases reported divergence in function, concluding that the balanced activity of these d , d ‐carboxypeptidases is required to ensure synthesis of a robust mature PG (Meiresonne, Ploeg, Hink, & Blaauwen, ).…”