Interrupting peptidoglycan deacetylation during Bdellovibrio predator-prey interaction prevents ultimate destruction of prey wall, liberating bacterial-ghosts.

Carey Lambert,Waldemar Vollmer,R Elizabeth Sockett,Nhat Khai Bui,Shin-Ichi Aizawa,Susan Liddell,Andrew L Lovering,Ana Clark,Thomas R Lerner,Hannah Somers

doi:10.1038/srep26010

Abstract

The peptidoglycan wall, located in the periplasm between the inner and outer membranes of the cell envelope in Gram-negative bacteria, maintains cell shape and endows osmotic robustness. Predatory Bdellovibrio bacteria invade the periplasm of other bacterial prey cells, usually crossing the peptidoglycan layer, forming transient structures called bdelloplasts within which the predators replicate. Prey peptidoglycan remains intact for several hours, but is modified and then degraded by escaping predators. Here we show predation is altered by deleting two Bdellovibrio N-acetylglucosamine (GlcNAc) deacetylases, one of which we show to have a unique two domain structure with a novel regulatory”plug”. Deleting the deacetylases limits peptidoglycan degradation and rounded prey cell “ghosts” persist after mutant-predator exit. Mutant predators can replicate unusually in the periplasmic region between the peptidoglycan wall and the outer membrane rather than between wall and inner-membrane, yet still obtain nutrients from the prey cytoplasm. Deleting two further genes encoding DacB/PBP4 family proteins, known to decrosslink and round prey peptidoglycan, results in a quadruple mutant Bdellovibrio which leaves prey-shaped ghosts upon predation. The resultant bacterial ghosts contain cytoplasmic membrane within bacteria-shaped peptidoglycan surrounded by outer membrane material which could have promise as “bacterial skeletons” for housing artificial chromosomes.

Highlights

The peptidoglycan wall, located in the periplasm between the inner and outer membranes of the cell envelope in Gram-negative bacteria, maintains cell shape and endows osmotic robustness
As transcription of the bd0468 and bd3279 genes peaked sharply at the point of prey-bacterial invasion by B. bacteriovorus HD100, but declined during predatory growth inside bacteria (Fig. 2), we reasoned that the gene products were deacetylating the prey peptidoglycan during early stages of predation
B. bacteriovorus ∆bd0468bd3279 was still predatory, progeny Bdellovibrio were released more slowly from prey bdelloplasts compared to wild type (Fig. 4) as determined by measuring the time from progeny maturation as the growing, long intraperiplasmic Bdellovibrio divides into multiple shorter progeny to the time that the first progeny Bdellovibrio leaves the outer layers of the prey

Summary

Introduction

The peptidoglycan wall, located in the periplasm between the inner and outer membranes of the cell envelope in Gram-negative bacteria, maintains cell shape and endows osmotic robustness. We discovered[1,2] that two predatory enzymes (Bd3459 and Bd0816) of the DacB/Penicillin-binding-protein (PBP) 4 family cause rounding of the Gram-negative prey cell walls ( called sacculi) and that this allowed rapid invasion of the prey-bacterial periplasm and establishment of an infected bacterial prey-bdelloplast with a growing Bdellovibrio inside. Such “hollowed out” bacterial ghosts can be prepared from diverse Gram-negative bacterial species and could have future applications in synthetic biology

Methods

Results

Conclusion