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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).

This family belongs to the: Major Facilitator (MFS) Superfamily.

References associated with 4.H.1 family:

Ernst, C.M. and A. Peschel. (2011). Broad-spectrum antimicrobial peptide resistance by MprF-mediated aminoacylation and flipping of phospholipids. Mol. Microbiol. 80: 290-299. 21306448
Ernst, C.M., C.J. Slavetinsky, S. Kuhn, J.N. Hauser, M. Nega, N.N. Mishra, C. Gekeler, A.S. Bayer, and A. Peschel. (2018). Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin Resistance. MBio 9:. 30563904
Ernst, C.M., S. Kuhn, C.J. Slavetinsky, B. Krismer, S. Heilbronner, C. Gekeler, D. Kraus, S. Wagner, and A. Peschel. (2015). The lipid-modifying multiple peptide resistance factor is an oligomer consisting of distinct interacting synthase and flippase subunits. MBio 6:. 25626904
Hebecker, S., J. Krausze, T. Hasenkampf, J. Schneider, M. Groenewold, J. Reichelt, D. Jahn, D.W. Heinz, and J. Moser. (2015). Structures of two bacterial resistance factors mediating tRNA-dependent aminoacylation of phosphatidylglycerol with lysine or alanine. Proc. Natl. Acad. Sci. USA 112: 10691-10696. 26261323
Helmstetter, F., P. Arnold, B. Höger, L.M. Petersen, and E. Beitz. (2019). Formate-nitrite transporters carrying nonprotonatable amide amino acids instead of a central histidine maintain pH-dependent transport. J. Biol. Chem. 294: 623-631. 30455351
Kristian, S.A., M. Dürr, J.A. Van Strijp, B. Neumeister, and A. Peschel. (2003). MprF-mediated lysinylation of phospholipids in Staphylococcus aureus leads to protection against oxygen-independent neutrophil killing. Infect. Immun. 71: 546-549. 12496209
Peschel, A., R.W. Jack, M. Otto, L.V. Collins, P. Staubitz, G. Nicholson, H. Kalbacher, W.F. Nieuwenhuizen, G. Jung, A. Tarkowski, K.P. van Kessel, and J.A. van Strijp. (2001). Staphylococcus aureus resistance to human defensins and evasion of neutrophil killing via the novel virulence factor MprF is based on modification of membrane lipids with l-lysine. J Exp Med 193: 1067-1076. 11342591
Slavetinsky, C.J., A. Peschel, and C.M. Ernst. (2012). Alanyl-phosphatidylglycerol and lysyl-phosphatidylglycerol are translocated by the same MprF flippases and have similar capacities to protect against the antibiotic daptomycin in Staphylococcus aureus. Antimicrob. Agents Chemother. 56: 3492-3497. 22491694