YhdP, TamB, and YdbH Are Redundant but Essential for Growth and Lipid Homeostasis of the Gram-Negative Outer Membrane

The bacterial cell envelope is the first line of defense and point of contact with the environment and other organisms. Envelope biogenesis is therefore crucial for the survival and physiology of bacteria and is often targeted by antimicrobials. Gram-negative bacteria have a multilayered envelope delimited by an inner and outer membrane (IM and OM, respectively). The OM is a barrier against many antimicrobials because of its asymmetric lipid structure, with phospholipids composing the inner leaflet and lipopolysaccharides (LPS) the outer leaflet. Since lipid synthesis occurs at the IM, phospholipids and LPS are transported across the cell envelope and asymmetrically assembled at the OM during growth. How phospholipids are transported to the OM remains unknown. Recently, the Escherichia coli protein YhdP has been proposed to participate in this process through an unknown mechanism. YhdP belongs to the AsmA-like clan and contains domains homologous to those found in lipid transporters. Here, we used genetics to investigate the six members of the AsmA-like clan of proteins in E. coli. Our data show that YhdP and its paralogs TamB and YdbH are redundant, but not equivalent, in performing an essential function in the cell envelope. Among the AsmA-like paralogs, only the combined loss of YhdP, TamB, and YdbH is lethal, and any of these three proteins is sufficient for growth. We also show that these proteins are required for OM lipid homeostasis and propose that they are the long-sought-after phospholipid transporters that are required for OM biogenesis.