The Gram-negative outer-membrane envelops the bacterium and functions as a permeability barrier against antibiotics, detergents, and environmental stresses. Some virulence factors serve to maintain the integrity of the outer membrane, including DolP (formerly YraP) a protein of unresolved structure and function. Here, we reveal DolP is a lipoprotein functionally conserved amongst Gram-negative bacteria and that loss of DolP increases membrane fluidity. We present the NMR solution structure for Escherichia coli DolP, which is composed of two BON domains that form an interconnected opposing pair. The C-terminal BON domain binds anionic phospholipids through an extensive membrane:protein interface. This interaction is essential for DolP function and is required for sub-cellular localisation of the protein to the cell division site, providing evidence of subcellular localisation of these phospholipids within the outer membrane. The structure of DolP provides a new target for developing therapies that disrupt the integrity of the bacterial cell envelope.

Biogenesis of the bacterial outer membrane is key to survival and antibiotic resistance. Central to this is the beta-barrel assembly machine (Bam) complex and its associated chaperones, which are responsible for outer membrane protein (OMP) transport and insertion. The Escherichia coli Bam complex consists of two essential subunits, BamA and BamD, and three non-essential lipoproteins, BamB, BamC and BamE. Optimal Bam function is further dependent on the non-essential chaperones DegP, Skp and SurA. Despite intensive study, the specific function of these non-essential Bam-associated proteins remains unknown. Here, we analysed knockout strains for each gene by phenotypic screening, conservation analysis and high-throughput genetics. We reveal that Bam activity is affected by outer membrane lipid composition and that enterobacterial common antigen is essential in the absence of the chaperone SurA. We also show that components of peptidoglycan are conditionally essential with Bam accessory lipoproteins and that DNA replication is perturbed in the absence of BamB. Together, our data indicates potential mechanisms for coordination of OMP biogenesis with processes such as LPS and peptidoglycan biogenesis, and DNA replication.

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