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bioRxiv · 10.1101/2025.04.02.646830

The aPBP-type cell wall synthase PBP1b plays a specialized role in fortifying the Escherichia coli division site against osmotic rupture

Navarro, P. P.·Vettiger, A.·Hajdu, R.·Ananda, V. Y.·Lopez-Tavares, A.·Schmid, E.·Walter, J.·Loose, M.·Chao, L. H.·Bernhardt, T. G.
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Abstract

A multi-protein system called the divisome promotes bacterial division. This apparatus synthesizes the peptidoglycan (PG) cell wall layer that forms the daughter cell poles and protects them from osmotic lysis. In the model Gram-negative bacterium Escherichia coli, PG synthases called class A penicillin-binding proteins (aPBPs) have been proposed to play crucial roles in division. However, there is limited experimental support for aPBPs playing a specialized role in division that is distinct from their general function in the expansion and fortification of the PG matrix. Here, we present in situ cryogenic electron tomography data indicating that the aPBP-type enzyme PBP1b is required to produce a wedge-like density of PG at the division site. Furthermore, atomic force and live cell microscopy showed that loss of this structure weakens the division site and renders it susceptible to lysis. Surprisingly, we found that the lipoprotein activator LpoB needed to promote the general function of PBP1b was not required for normal division site architecture or its integrity. Additionally, we show that of the two PBP1b isoforms produced in cells, it is the one with an extended cytoplasmic N-terminus that functions in division, likely via recruitment by the FtsA component of the divisome. Altogether, our results demonstrate that PBP1b plays a specialized, LpoB-independent role in E. coli cell division involving the biogenesis of a PG structure that prevents osmotic rupture. The conservation of aPBPs with extended cytoplasmic N-termini suggests that other Gram-negative bacteria may use similar mechanisms to reinforce their division site.

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Navarro, P. P., Vettiger, A., Hajdu, R., Ananda, V. Y., Lopez-Tavares, A., Schmid, E., Walter, J., Loose, M., Chao, L. H., Bernhardt, T. G.. 2025-04-02. The aPBP-type cell wall synthase PBP1b plays a specialized role in fortifying the Escherichia coli division site against osmotic rupture. https://doi.org/10.1101/2025.04.02.646830

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