Evidence for phospholipid export from the bacterial inner membrane by the Mla ABC transport system

Abstract

The Mla pathway is believed to be involved in maintaining the asymmetrical Gram-negative outer membrane via retrograde phospholipid transport. The pathway is composed of three components: the outer membrane MlaA–OmpC/F complex, a soluble periplasmic protein, MlaC, and the inner membrane ATPase, MlaFEDB complex. Here, we solve the crystal structure of MlaC in its phospholipid-free closed apo conformation, revealing a pivoting β-sheet mechanism that functions to open and close the phospholipid-binding pocket. Using the apo form of MlaC, we provide evidence that the inner-membrane MlaFEDB machinery exports phospholipids to MlaC in the periplasm. Furthermore, we confirm that the phospholipid export process occurs through the MlaD component of the MlaFEDB complex and that this process is independent of ATP. Our data provide evidence of an apparatus for lipid export away from the inner membrane and suggest that the Mla pathway may have a role in anterograde phospholipid transport.

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Fig. 1: The MlaC-apo structure.
Fig. 2: MlaC requires MlaD for PL loading.
Fig. 3: MlaFEDB is active and exports IM PLs to MlaC.
Fig. 4: QCM–D and ATR-FTIR further suggest MlaFEDB exports IM PLs to MlaC.
Fig. 5: MlaFEDB-driven PL export occurs in an ATPase-independent manner but ATP hydrolysis is linked to MlaC-apo binding.
Fig. 6: Schematic summarizing the questions for the mechanisms of the Mla system arising from this study.

Data availability

The MlaC-apo X-ray structure has been deposited in the PDB with the accession number 6GKI. In addition, the data that support the findings of this study are available from the corresponding author on request.

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Acknowledgements

We thank D. Ekiert and G. Bhabha for their discussions and for providing the Mla plasmid constructs. We thank the HWB-NMR at the University of Birmingham for providing open access to their Wellcome Trust-funded NMR equipment. This research was supported by the BBSRC grant no. BB/P009840/1 (T.J.K. and G.W.H.) and Wellcome Trust grant no. 208400/Z/17/Z. T.J.P. acknowledges the use of the Iridis high-performance computing resources at the University of Southampton.

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G.W.H., S.C.L.H., C.S.L., P.S., C.H., T.J.P., P.J.W., A.C.L., D.G.W., M.J., V.S., I.T.C., C.H., G.L.I., J.A.B., Y.Y., M.J., D.H., I.R.H., L.A.C., A.L.L. and T.J.K. participated in the conception and design of the work. G.W.H., S.C.L.H., C.S.L., P.S., A.H.M., T.J.P., P.J.W., A.C.L., D.G.W., R.J.P., L.A.C., A.L.L. and T.J.K. participated in the data acquisition, analysis or interpretation of the work. G.W.H., S.C.L.H., C.S.L., P.S., T.J.P., P.J.W., A.C.L., D.G.W., L.A.C., A.L.L. and T.J.K. were involved in writing and editing the manuscript. All authors approved the final version submitted for publication.

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Correspondence to Timothy J. Knowles.

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Hughes, G.W., Hall, S.C.L., Laxton, C.S. et al. Evidence for phospholipid export from the bacterial inner membrane by the Mla ABC transport system. Nat Microbiol 4, 1692–1705 (2019). https://doi.org/10.1038/s41564-019-0481-y

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