Structure of the MlaC-MlaD complex reveals molecular basis of periplasmic phospholipid transport.
Journal
Nature communications
ISSN: 2041-1723
Titre abrégé: Nat Commun
Pays: England
ID NLM: 101528555
Informations de publication
Date de publication:
30 Jul 2024
30 Jul 2024
Historique:
received:
06
04
2023
accepted:
08
07
2024
medline:
31
7
2024
pubmed:
31
7
2024
entrez:
30
7
2024
Statut:
epublish
Résumé
The Maintenance of Lipid Asymmetry (Mla) pathway is a multicomponent system found in all gram-negative bacteria that contributes to virulence, vesicle blebbing and preservation of the outer membrane barrier function. It acts by removing ectopic lipids from the outer leaflet of the outer membrane and returning them to the inner membrane through three proteinaceous assemblies: the MlaA-OmpC complex, situated within the outer membrane; the periplasmic phospholipid shuttle protein, MlaC; and the inner membrane ABC transporter complex, MlaFEDB, proposed to be the founding member of a structurally distinct ABC superfamily. While the function of each component is well established, how phospholipids are exchanged between components remains unknown. This stands as a major roadblock in our understanding of the function of the pathway, and in particular, the role of ATPase activity of MlaFEDB is not clear. Here, we report the structure of E. coli MlaC in complex with the MlaD hexamer in two distinct stoichiometries. Utilising in vivo complementation assays, an in vitro fluorescence-based transport assay, and molecular dynamics simulations, we confirm key residues, identifying the MlaD β6-β7 loop as essential for MlaCD function. We also provide evidence that phospholipids pass between the C-terminal helices of the MlaD hexamer to reach the central pore, providing insight into the trajectory of GPL transfer between MlaC and MlaD.
Identifiants
pubmed: 39080293
doi: 10.1038/s41467-024-50615-3
pii: 10.1038/s41467-024-50615-3
doi:
Substances chimiques
Phospholipids
0
Escherichia coli Proteins
0
ATP-Binding Cassette Transporters
0
MlaD protein, E coli
0
Phospholipid Transfer Proteins
0
Membrane Proteins
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
6394Subventions
Organisme : RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
ID : BB/S017283/1
Organisme : RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
ID : BB/M01116X/1
Organisme : RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
ID : BB/M01116X/1
Organisme : RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
ID : BB/M01116X/1
Organisme : RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
ID : BB/S017283/1
Organisme : RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
ID : BB/M01116X/1
Organisme : RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
ID : BB/M01116X/1
Organisme : RCUK | Biotechnology and Biological Sciences Research Council (BBSRC)
ID : BB/M01116X/1
Informations de copyright
© 2024. The Author(s).
Références
Nikaido, H. Restoring permeability barrier function to outer membrane. Chem. Biol. 12, 507–509 (2005).
doi: 10.1016/j.chembiol.2005.05.001
pubmed: 15911368
Sperandeo, P., Martorana, A. M. & Polissi, A. Lipopolysaccharide biogenesis and transport at the outer membrane of Gram-negative bacteria. Biochim. Biophys. Acta Mol. Cell Biol. Lipids 1862, 1451–1460 (2017).
doi: 10.1016/j.bbalip.2016.10.006
pubmed: 27760389
Powers, M. J. & Trent, M. S. Intermembrane transport: glycerophospholipid homeostasis of the Gram-negative cell envelope. Proc. Natl Acad. Sci. USA 116, 17147–17155 (2019).
doi: 10.1073/pnas.1902026116
pubmed: 31420510
pmcid: 6717313
Low, W. Y., Thong, S. & Chng, S. S. ATP disrupts lipid-binding equilibrium to drive retrograde transport critical for bacterial outer membrane asymmetry. Proc. Natl Acad. Sci. USA 118, e2110055118 (2021).
doi: 10.1073/pnas.2110055118
pubmed: 34873038
pmcid: 8685716
Giacometti, S. I., MacRae, M. R., Dancel-Manning, K., Bhabha, G. & Ekiert, D. C. Lipid Transport Across Bacterial Membranes. Annu. Rev. Cell Dev. Biol. 38, 125–153 (2022).
doi: 10.1146/annurev-cellbio-120420-022914
pubmed: 35850151
Malinverni, J. C. & Silhavy, T. J. An ABC transport system that maintains lipid asymmetry in the gram-negative outer membrane. Proc. Natl Acad. Sci. USA 106, 8009–8014 (2009).
doi: 10.1073/pnas.0903229106
pubmed: 19383799
pmcid: 2683108
Chong, Z. S., Woo, W. F. & Chng, S. S. Osmoporin OmpC forms a complex with MlaA to maintain outer membrane lipid asymmetry in Escherichia coli. Mol. Microbiol. 98, 1133–1146 (2015).
doi: 10.1111/mmi.13202
pubmed: 26314242
Abellon-Ruiz, J. et al. Structural basis for maintenance of bacterial outer membrane lipid asymmetry. Nat. Microbiol. 2, 1616–1623 (2017).
doi: 10.1038/s41564-017-0046-x
pubmed: 29038444
Hughes, G. W. et al. Evidence for phospholipid export from the bacterial inner membrane by the Mla ABC transport system. Nat. Microbiol. 4, 1692–1705 (2019).
doi: 10.1038/s41564-019-0481-y
pubmed: 31235958
Thong, S. et al. Defining key roles for auxiliary proteins in an ABC transporter that maintains bacterial outer membrane lipid asymmetry. eLife 5, e19042 (2016).
doi: 10.7554/eLife.19042
pubmed: 27529189
pmcid: 5016091
Kolich, L. R. et al. Structure of MlaFB uncovers novel mechanisms of ABC transporter regulation. eLife 9, e60030 (2020).
doi: 10.7554/eLife.60030
pubmed: 32602838
pmcid: 7367683
Chi, X. et al. Structural mechanism of phospholipids translocation by MlaFEDB complex. Cell Res. 30, 1127–1135 (2020).
doi: 10.1038/s41422-020-00404-6
pubmed: 32884137
pmcid: 7784689
Zhang, Y., Fan, Q., Chi, X., Zhou, Q. & Li, Y. Cryo-EM structures of Acinetobacter baumannii glycerophospholipid transporter. Cell Discov. 6, 86 (2020).
doi: 10.1038/s41421-020-00230-5
pubmed: 33298869
pmcid: 7677376
Mann, D. et al. Structure and lipid dynamics in the maintenance of lipid asymmetry inner membrane complex of A. baumannii. Commun. Biol. 4, 817 (2021).
doi: 10.1038/s42003-021-02318-4
pubmed: 34188171
pmcid: 8241846
Tang, X. et al. Structural insights into outer membrane asymmetry maintenance in Gram-negative bacteria by MlaFEDB. Nat. Struct. Mol. Biol. 28, 81–91 (2021).
doi: 10.1038/s41594-020-00532-y
pubmed: 33199922
Zhou, C. et al. Structural insight into phospholipid transport by the MlaFEBD complex from P. aeruginosa. J. Mol. Biol. 433, 166986 (2021).
doi: 10.1016/j.jmb.2021.166986
pubmed: 33845086
Coudray, N., Isom, G. L., MacRae, M. R., Saiduddin, M. N., Bhabha, G. & Ekiert, D. C. Structure of bacterial phospholipid transporter MlaFEDB with substrate bound. eLife 9, e62518 (2020).
doi: 10.7554/eLife.62518
pubmed: 33236984
pmcid: 7790496
Ekiert, D. C. et al. Architectures of lipid transport systems for the bacterial outer membrane. Cell 169, 273–285.e217 (2017).
doi: 10.1016/j.cell.2017.03.019
pubmed: 28388411
pmcid: 5467742
Ercan, B., Low, W. Y., Liu, X. & Chng, S. S. Characterization of interactions and phospholipid transfer between substrate binding proteins of the OmpC-Mla system. Biochemistry 58, 114–119 (2018).
MacRae, M. R. et al. Protein-protein interactions in the Mla lipid transport system probed by computational structure prediction and deep mutational scanning. J. Biol. Chem. 299, 104744 (2023).
doi: 10.1016/j.jbc.2023.104744
pubmed: 37100290
pmcid: 10245069
Jumper, J. et al. Highly accurate protein structure prediction with AlphaFold. Nature 596, 583–589 (2021).
doi: 10.1038/s41586-021-03819-2
pubmed: 34265844
pmcid: 8371605
Jones, N. C. & Osborn, M. J. Translocation of phospholipids between the outer and inner membranes of Salmonella typhimurium. J. Biol. Chem. 252, 7405–7412 (1977).
doi: 10.1016/S0021-9258(19)66979-6
pubmed: 332696
Donohue-Rolfe, A. M. & Schaechter, M. Translocation of phospholipids from the inner to the outer membrane of Escherichia coli. Proc. Natl Acad. Sci. USA 77, 1867–1871 (1980).
doi: 10.1073/pnas.77.4.1867
pubmed: 6246510
pmcid: 348609
Langley, K. E., Hawrot, E. & Kennedy, E. P. Membrane assembly: movement of phosphatidylserine between the cytoplasmic and outer membranes of Escherichia coli. J. Bacteriol. 152, 1033–1041 (1982).
doi: 10.1128/jb.152.3.1033-1041.1982
pubmed: 6754694
pmcid: 221606
Szollosi, D., Rose-Sperling, D., Hellmich, U. A. & Stockner, T. Comparison of mechanistic transport cycle models of ABC exporters. Biochim. Biophys. Acta Biomembr. 1860, 818–832 (2018).
doi: 10.1016/j.bbamem.2017.10.028
pubmed: 29097275
Thomason, L. C., Costantino, N. & Court, D. L. E. coli genome manipulation by P1 transduction. Curr. Protoc. Mol. Biol. 1, 1 17 11–11 17 18 (2007).
Datsenko, K. A. & Wanner, B. L. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc. Natl Acad. Sci. USA 97, 6640–6645 (2000).
doi: 10.1073/pnas.120163297
pubmed: 10829079
pmcid: 18686
Baba, T. et al. Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol. Syst. Biol. 2, 2006 0008 (2006).
doi: 10.1038/msb4100050
pubmed: 16738554
pmcid: 1681482
Goddard, T. D. et al. UCSF ChimeraX: meeting modern challenges in visualization and analysis. Protein Sci. 27, 14–25 (2018).
doi: 10.1002/pro.3235
pubmed: 28710774
Pettersen, E. F. et al. UCSF ChimeraX: structure visualization for researchers, educators, and developers. Protein Sci. 30, 70–82 (2021).
doi: 10.1002/pro.3943
pubmed: 32881101
Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. Sect. D., Biol. Crystallogr. 60, 2126–2132 (2004).
doi: 10.1107/S0907444904019158
Liebschner, D. et al. Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Acta Crystallogr D. Struct. Biol. 75, 861–877 (2019).
doi: 10.1107/S2059798319011471
pubmed: 31588918
pmcid: 6778852
Schrödinger. The PyMOL Molecular Graphics System, Version 20 Schrödinger, LLC (2019).
Abraham, M. J. et al. GROMACS: high performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX 1–2, 19–25 (2015).
doi: 10.1016/j.softx.2015.06.001
Stansfeld, P. J. et al. MemProtMD: automated insertion of membrane protein structures into explicit lipid membranes. Structure 23, 1350–1361 (2015).
doi: 10.1016/j.str.2015.05.006
pubmed: 26073602
pmcid: 4509712
Nugent, T. & Jones, D. T. Membrane protein orientation and refinement using a knowledge-based statistical potential. BMC Bioinform. 14, 276 (2013).
doi: 10.1186/1471-2105-14-276
Souza, P. C. T. et al. Martini 3: a general purpose force field for coarse-grained molecular dynamics. Nat. Methods 18, 382–388 (2021).
doi: 10.1038/s41592-021-01098-3
pubmed: 33782607
Wassenaar, T. A., Ingolfsson, H. I., Bockmann, R. A., Tieleman, D. P. & Marrink, S. J. Computational lipidomics with insane: a versatile tool for generating custom membranes for molecular simulations. J. Chem. Theory Comput. 11, 2144–2155 (2015).
doi: 10.1021/acs.jctc.5b00209
pubmed: 26574417
Hess, B., Bekker, H., Berendsen, H. J. & Fraaije, J. G. E. M. LINCS: a linear constraint solver for molecular simulations. J. Comput. Chem. 18, 10 (1998).
Stansfeld, P. J. & Sansom, M. S. From coarse grained to atomistic: a serial multiscale approach to membrane protein simulations. J. Chem. Theory Comput. 7, 1157–1166 (2011).
doi: 10.1021/ct100569y
pubmed: 26606363
Vickery, O. N. & Stansfeld, P. J. CG2AT2: an enhanced fragment-based approach for serial multi-scale molecular dynamics simulations. J. Chem. Theory Comput. 17, 6472–6482 (2021).
doi: 10.1021/acs.jctc.1c00295
pubmed: 34492188
pmcid: 8515810