Skip to main page content
U.S. flag

An official website of the United States government

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2020 Apr 7;28(4):475-487.e3.
doi: 10.1016/j.str.2020.01.012. Epub 2020 Feb 12.

Characterizing Membrane Association and Periplasmic Transfer of Bacterial Lipoproteins through Molecular Dynamics Simulations

Affiliations

Characterizing Membrane Association and Periplasmic Transfer of Bacterial Lipoproteins through Molecular Dynamics Simulations

Shanlin Rao et al. Structure. .

Abstract

Escherichia coli lipoprotein precursors at the inner membrane undergo three maturation stages before transport by the Lol system to the outer membrane. Here, we develop a pipeline to simulate the membrane association of bacterial lipoproteins in their four maturation states. This has enabled us to model and simulate 81 of the predicted 114 E. coli lipoproteins and reveal their interactions with the host lipid membrane. As part of this set we characterize the membrane contacts of LolB, the lipoprotein involved in periplasmic translocation. We also consider the means and bioenergetics for lipoprotein localization. Our calculations uncover a preference for LolB over LolA and therefore indicate how a lipoprotein may be favorably transferred from the inner to outer membrane. Finally, we reveal that LolC has a role in membrane destabilization, thereby promoting lipoprotein transfer to LolA.

Keywords: antibiotics; biogenesis; lipoprotein; membrane proteins; molecular simulation.

PubMed Disclaimer

Conflict of interest statement

Declaration of Interests The authors declare no competing interests.

Figures

None
Graphical abstract
Figure 1
Figure 1
The Lipoprotein Biogenesis Pathway Sec-secreted E. coli lipoprotein precursors (preprolipoproteins) are sequentially post-translationally modified in the IM, by preprolipoprotein diacylglyceryl transferase (Lgt), prolipoprotein signal peptidase (LspA), and apolipoprotein N-acyltransferase (Lnt). Upon maturation, lipoproteins are either retained in the IM or transported by the Lol machinery to the inner leaflet of the OM. The Lol system is comprised of an ATP-binding cassette transporter (LolCDE), a periplasmic carrier protein LolA, and an OM receptor protein LolB that is itself a lipoprotein. Example lipoproteins Pal, BamBCDE, and LptE are shown.
Figure 2
Figure 2
Workflow for Modeling and Simulating the Four Stages of Bacterial Lipoprotein Biogenesis (A) The structure of a lipoprotein is reoriented so that the N-terminal end is positioned closest to the membrane. A flexible tether is modeled onto the structure, with SP, SP and diacylcysteine (SP + Di), diacylcysteine (Di), or triacylcysteine (Tri) modification. The states were subjected to CG-MD simulations. (B) CG-MD snapshots for a subset of the 30 molecular structures of E. coli lipoproteins. The membrane association with the four discrete membrane anchors is shown for each lipoprotein. In each case, the membrane association of the lipoprotein is shown at the end of a 1 μs simulation, with either its SP (top left), SP + Di (top right), Di (bottom left), or Tri modification (bottom right). (C) Membrane localization of the conserved cysteine residue in the four stages of maturation.
Figure 3
Figure 3
Membrane Associations of Lipoproteins (A) Exemplar membrane interactions of the 81 triacyl-modified lipoproteins bound to a model IM, before their translocation across the periplasm to the OM. (B) Residue-lipid contacts for bacterial lipoproteins. Assessment of the residue interactions with the glycerol group of POPG (GL0), ethanolamine group of POPE (NH3), phosphate groups (PO4), glycerol (GL1:2), and acyl tails (Tail) of both lipids.
Figure 4
Figure 4
Reproducibility of Membrane Association of the 81 Modeled Triacylated E. coli Lipoproteins For each of the 81 modeled lipoprotein structures the similarity in the membrane-protein interactions are compared in the five repeats of membrane association. A dark blue color reflects faithful reproduction in binding mode between two simulations and a Pearson correlation coefficient of 1, while a red color highlights a distinct binding orientation and a Pearson correlation coefficient of -1 for the lipid-residue interactions. The five simulations per lipoprotein are labeled 1 through 5.
Figure 5
Figure 5
Molecular Simulations of Lipoprotein Complexes The methodology allows for the construction of lipoprotein tethers within macromolecular complexes; here shown for ten structures of E. coli K12 lipoprotein complexes, CusC, Lpp, CsgG, LptDE, BamABCDE, and NlpE in a model OM, MetQNI, CyoABC, and ApbE in a model IM, and the AcrABZ-TolC complex spanning the periplasm and inserted into both IM and OM. In each case the triacyl lipoprotein tether is shown in blue sticks, with the lipoprotein colored. The non-lipoprotein subunits are shown in gray.
Figure 6
Figure 6
Lipoproteins Involved in Periplasmic Transport Membrane association of (A) LolB, (B) MlaA, and (C) PqiC in their (i) primary and (ii) secondary binding orientations. Phosphate atoms are shown as red spheres. Proteins are shown as a cartoon representation, and colored on a white to blue scale, with blue indicating extensive lipid contacts. See also Supplemental Information, Figure S5.
Figure 7
Figure 7
Calculating the Energetics of Lipoprotein Transport The energetics associated with lipoprotein transfer across the periplasm, obtained from umbrella sampling and PMF calculations, calculated using WHAM with errors computed using Bayesian bootstrapping. Plots are shown for the extraction of the triacylated cysteine moiety from (A) IM, (B) LolA (orange), (C) LolB (cyan), and (D) OM. The depicted energy values are derived from the minimum value of the PMF.
Figure 8
Figure 8
Molecular Simulations of the Lol and Mac Transporter Complexes In the resting state, LolCDE (A) and LolACDE (B) complexes reveal extensive membrane deformation about the periplasmic domain of LolC, with the position of the phosphates changing by up to 20 Å from their bulk membrane position. The equivalent state of MacB (C) does not show membrane deformation, nor do simulations in the active, ATP-bound states of (D) LolCDE, (E) LolACDE, or (F) MacB. Proteins are shown in a cartoon representation, highlighting LolC (green), LolD (purple), LolE (yellow), LolA (orange), and MacB (gray). Cumulative phosphate positions from the simulations are shown as a surface, on a red-white-blue scale, from thickening to thinning of the membrane.
Figure 9
Figure 9
Thermodynamics of Lipoprotein Transfer (A) Structural basis for transport of triacylated LolB (blue)—itself a lipoprotein—from the IM to OM via LolA (orange) and LolB (cyan). (B) Thermodynamic cycle of lipoprotein transport, combining the values from the PMF calculations in Figure 7. The energy needed for membrane extraction is obtained from ATP binding and hydrolysis by LolCDE. Once extracted from the IM the bioenergetics of transport is downhill from LolA to LolB to the OM.

Similar articles

Cited by

References

    1. Abellon-Ruiz J., Kaptan S.S., Basle A., Claudi B., Bumann D., Kleinekathofer U., van den Berg B. Structural basis for maintenance of bacterial outer membrane lipid asymmetry. Nat. Microbiol. 2017;2:1616–1623. - PubMed
    1. Abraham M.J., Murtola T., Schulz R., Páll S., Smith J.C., Hess B., Lindahl E. GROMACS: high performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX. 2015;1-2:19–25.
    1. Abramson J., Riistama S., Larsson G., Jasaitis A., Svensson-Ek M., Laakkonen L., Puustinen A., Iwata S., Wikstrom M. The structure of the ubiquinol oxidase from Escherichia coli and its ubiquinone binding site. Nat. Struct. Biol. 2000;7:910–917. - PubMed
    1. Arnarez C., Marrink S.J., Periole X. Identification of cardiolipin binding sites on cytochrome c oxidase at the entrance of proton channels. Sci. Rep. 2013;3:1263. - PMC - PubMed
    1. Arnarez C., Mazat J.P., Elezgaray J., Marrink S.J., Periole X. Evidence for cardiolipin binding sites on the membrane-exposed surface of the cytochrome bc1. J. Am. Chem. Soc. 2013;135:3112–3120. - PubMed

Publication types

MeSH terms

LinkOut - more resources