4.H.1  The Lysyl Phosphatidylglycerol Synthase/Flippase (MprF) Family 

Phospholipids are synthesized at the inner leaflet of the bacterial cytoplasmic membrane but have to be translocated to the outer leaflet to maintain membrane lipid bilayer composition and structure. MprF is a large integral membrane protein found in several prokaryotic phyla, the C-terminus of which modifies phosphatidylglycerol (PG) with lysine or alanine to modulate the membrane surface charge and, as a consequence, confer resistance to cationic antimicrobial agents such as daptomycin (Ernst and Peschel 2011). In addition, MprF is a flippase for the resulting lipids, Lys-PG or Ala-PG. Ernst et al. 2015 demonstrated that the flippase activity resides in the N-terminal 6 to 8 transmembrane segments of the Staphylococcus aureus MprF and that several conserved, charged amino acids and a proline residue are crucial for flippase function. MprF protects S. aureus against the membrane-active antibiotic daptomycin only when both domains are present, but the two parts do not need to be covalently linked and can function in trans (Hebecker et al. 2015). The Lys-PG synthase and flippase domains were each found to homo-oligomerize and also to interact with each other, which illustrates how the two functional domains may act together. Moreover, full-length MprF proteins formed oligomers, indicating that MprF functions as a dimer or larger oligomer (Ernst et al. 2015). The data reveal how bacterial phospholipid flippases may function in the context of lipid biosynthetic processes. The coupling of lysyl transfer to lipid export is hypothetical.

Bacterial cytoplasmic membranes have to cope with membrane-damaging agents such as cationic antimicrobial peptides (CAMPs) produced, for example, by competing bacteria (bacteriocins), secreted by eukaryotic host cells (defensins), or used as antimicrobial therapy (daptomycin). MprF proteins are found in many Gram-positive, Gram-negative, and archaeal commensals or pathogens and confers resistance to CAMPs by modifying anionic phospholipids with amino acids, thereby compromising the membrane interaction of CAMPs. Ernst et al. 2015 described how MprF does not only modify phospholipids but uses an additional, distinct domain for translocating the resulting lysinylated phospholipids to the outer leaflet of the membrane. They revealed critical details for the structure and function of MprF (Ernst et al. 2015).  Moreover, gain-of-function mutations in MprF confer specific daptomycin resistance (Ernst et al. 2018).