The hit-and-run of cell wall synthesis: LpoB transiently binds and activates PBP1b through a conserved allosteric switch

• Published in: Nature communications Vol. 16; no. 1; pp. 6723 - 15
• Main Authors: Shlosman, Irina, Vettiger, Andrea, Bernhardt, Thomas G., Kruse, Andrew C., Loparo, Joseph J.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 21.07.2025; Nature Publishing Group; Nature Portfolio
• Subjects: 14/63; 631/326/41/2528; 631/326/41/2536; 631/45/607/1169; 631/57/2265; Allosteric properties; Allosteric Regulation; Antibiotics; Bacteria; Bacterial Outer Membrane Proteins - genetics; Bacterial Outer Membrane Proteins - metabolism; Bioengineering; Cell Wall - metabolism; Cell walls; Datasets; E coli; Efficiency; Enzymes; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Fluorescence Resonance Energy Transfer; Gram-negative bacteria; Humanities and Social Sciences; Lipids; Lipoproteins; Lipoproteins - metabolism; Molecular modelling; multidisciplinary; Mutation; Particle tracking; Penicillin; Penicillin-Binding Proteins - chemistry; Penicillin-Binding Proteins - genetics; Penicillin-Binding Proteins - metabolism; Peptidoglycan - biosynthesis; Peptidoglycan - metabolism; Peptidoglycan Glycosyltransferase; Peptidoglycans; Protein Binding; Proteins; Science; Science (multidisciplinary); Serine-Type D-Ala-D-Ala Carboxypeptidase - chemistry; Serine-Type D-Ala-D-Ala Carboxypeptidase - genetics; Serine-Type D-Ala-D-Ala Carboxypeptidase - metabolism; Synthesis
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-025-62051-y

The peptidoglycan (PG) cell wall is the primary protective layer of bacteria, making the process of PG synthesis a key antibiotic target. Class A penicillin-binding proteins (aPBPs) are a family of conserved and ubiquitous PG synthases that fortify and repair the PG matrix. In gram-negative bacteria, these enzymes are regulated by outer-membrane tethered lipoproteins. However, the molecular mechanism by which lipoproteins coordinate the spatial recruitment and enzymatic activation of aPBPs remains unclear. Here we use single-molecule FRET and single-particle tracking in E. coli to show that a prototypical lipoprotein activator LpoB triggers site-specific PG synthesis by PBP1b through conformational rearrangements. Once synthesis is initiated, LpoB affinity for PBP1b dramatically decreases and it dissociates from the synthesizing enzyme. Our results suggest that transient allosteric coupling between PBP1b and LpoB directs PG synthesis to areas of low peptidoglycan density, while simultaneously facilitating efficient lipoprotein redistribution to other sites in need of fortification. Class A PBPs accomplish cell wall synthesis to enable bacterial growth and are subject to inhibition by penicillin-type antibiotics. Here, authors leverage single-molecule and bioengineering approaches to show how these essential enzymes are regulated by cognate lipoprotein cofactors.