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Open AccessOpen Access JUSTC Chemistry 17 January 2024

Molecular mechanism underlying ABC exporter gating: a computational study

  • Department of Chemical Physics, University of Science and Technology of China, Hefei 230026, China

Cite this:
https://doi.org/10.52396/JUSTC-2022-0134
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  • Author Bio:

    Zi Wang received his Ph.D. degree in Chemistry from the University of Science and Technology of China. He is currently a senior engineer in biochemical research and development at the BGI Research, Shenzhen. His research mainly focuses on protein design and directed evolution of enzymes

    Jielou Liao is a Professor in Physical Chemistry at the University of Science and Technology of China. His current research interests include theoretical and computational biology and chemistry as well as quantum biology

  • Corresponding author: E-mail: liaojl@ustc.edu.cn
  • Received Date: 01 October 2022
  • Accepted Date: 04 November 2022
    • Available Online: 17 January 2024
    Abstract

  • Abstract

    ATP-binding cassette (ABC) exporters are a class of molecular machines that transport substrates out of biological membranes by gating movements leading to transitions between outward-facing (OF) and inward-facing (IF) conformational states. Despite significant advances in structural and functional studies, the molecular mechanism underlying conformational gating in ABC exporters is not completely understood. A complete elucidation of the state transitions during the transport cycle is beyond the capability of the all-atom molecular dynamics (MD) method because of the limited time scale of MD. In the present work, a coarse-grained molecular dynamics (CG-MD) method with an improved sampling strategy is performed for the bacterial ABC exporter MsbA. The resultant potential of the mean force (PMF) along the center-of-mass (COM) distances, d1 and d2, between the two opposing subunits of the internal and external gates, respectively, are obtained, delicately showing the details of the $ {\rm{OF}}\to {\rm{IF}} $ transition occurring via an occluded (OC) state, in which the internal and external gates are both closed. The OC state has an important role in the unidirectionality of the transport function of ABC exporters. Our CG-MD simulations dynamically show that upon NBD dissociation, the opening of the internal gate occurs in a highly cooperative manner with the closure of the external gate. Based on our PMF calculations and CG-MD simulations in this paper, we proposed a mechanistic model that is significantly different from those recently published in the literature, shedding light on the molecular mechanism by which the ABC exporter executes conformational gating for substrate translocation.

    Graphical abstract

    Nonequilibrium coarse-grained (CG) molecular dynamics (MD) simulations show that the ATP-binding cassette (ABC) exporter performs highly cooperative gating movements during the conformational transitions.

    Abstract

    ATP-binding cassette (ABC) exporters are a class of molecular machines that transport substrates out of biological membranes by gating movements leading to transitions between outward-facing (OF) and inward-facing (IF) conformational states. Despite significant advances in structural and functional studies, the molecular mechanism underlying conformational gating in ABC exporters is not completely understood. A complete elucidation of the state transitions during the transport cycle is beyond the capability of the all-atom molecular dynamics (MD) method because of the limited time scale of MD. In the present work, a coarse-grained molecular dynamics (CG-MD) method with an improved sampling strategy is performed for the bacterial ABC exporter MsbA. The resultant potential of the mean force (PMF) along the center-of-mass (COM) distances, d1 and d2, between the two opposing subunits of the internal and external gates, respectively, are obtained, delicately showing the details of the $ {\rm{OF}}\to {\rm{IF}} $ transition occurring via an occluded (OC) state, in which the internal and external gates are both closed. The OC state has an important role in the unidirectionality of the transport function of ABC exporters. Our CG-MD simulations dynamically show that upon NBD dissociation, the opening of the internal gate occurs in a highly cooperative manner with the closure of the external gate. Based on our PMF calculations and CG-MD simulations in this paper, we proposed a mechanistic model that is significantly different from those recently published in the literature, shedding light on the molecular mechanism by which the ABC exporter executes conformational gating for substrate translocation.

    • Public Summary

    • The CG-MD trajectory discloses highly cooperative gating movements in the MsbA ABC exporter protein.
    • The potential mean force (PMF) is used to capture the conformational transitions between the OF, OC and IF states, and thus, a detailed understanding of the mechanism of ABC exporter gating is achieved at a molecular level.
    • On the basis of the CG-MD simulation results, a mechanistic model, which is significantly different from those published in the literature, is proposed.

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    • References(31)
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    Thomas C, Tampé R. Structural and mechanistic principles of ABC transporters. Annu. Rev. Biochem., 2020, 89: 605–636. doi: 10.1146/annurev-biochem-011520-105201
    [2]
    Srikant S, Gaudet R. Mechanics and pharmacology of substrate selection and transport by eukaryotic ABC exporters. Nat. Struct. Mol. Biol., 2019, 26 (9): 792–801. doi: 10.1038/s41594-019-0280-4
    [3]
    Hofmann S, Januliene D, Mehdipour A R, et al. Conformation space of a heterodimeric ABC exporter under turnover conditions. Nature, 2019, 571 (7766): 580–583. doi: 10.1038/s41586-019-1391-0
    [4]
    Locher K P. Mechanistic diversity in ATP-binding cassette (ABC) transporters. Nat. Struct. Mol. Biol., 2016, 23 (6): 487–493. doi: 10.1038/nsmb.3216
    [5]
    Robey R W, Pluchino K M, Hall M D, et al. Revisiting the role of ABC transporters in multidrug-resistant cancer. Nat. Rev. Cancer, 2018, 18 (7): 452–464. doi: 10.1038/s41568-018-0005-8
    [6]
    Dawson R J P, Locher K P. Structure of a bacterial multidrug ABC transporter. Nature, 2006, 443 (7108): 180–185. doi: 10.1038/nature05155
    [7]
    Dawson R J P, Locher K P. Structure of the multidrug ABC transporter Sav1866 from Staphylococcus aureus in complex with AMP-PNP. FEBS Lett., 2007, 581 (5): 935–938. doi: 10.1016/j.febslet.2007.01.073
    [8]
    Ward A, Reyes C L, Yu J, et al. Flexibility in the ABC transporter MsbA: Alternating access with a twist. Proc. Natl. Acad. Sci. U.S.A., 2007, 104 (48): 19005–19010. doi: 10.1073/pnas.0709388104
    [9]
    Mi W, Li Y, Yoon S H, et al. Structural basis of MsbA-mediated lipopolysaccharide transport. Nature, 2017, 549 (7671): 233–237. doi: 10.1038/nature23649
    [10]
    Ho H, Miu A, Alexander M K, et al. Structural basis for dual-mode inhibition of the ABC transporter MsbA. Nature, 2018, 557 (7704): 196–201. doi: 10.1038/s41586-018-0083-5
    [11]
    Padayatti P S, Lee S C, Stanfield R L, et al. Structural insights into the lipid A transport pathway in MsbA. Structure, 2019, 27 (7): 1114–1123.e3. doi: 10.1016/j.str.2019.04.007
    [12]
    Angiulli G, Dhupar H S, Suzuki H, et al. New approach for membrane protein reconstitution into peptidiscs and basis for their adaptability to different proteins. eLife, 2020, 9: e53530. doi: 10.7554/eLife.53530
    [13]
    Thélot F A, Zhang W, Song K, et al. Distinct allosteric mechanisms of first-generation MsbA inhibitors. Science, 2021, 374 (6567): 580–585. doi: 10.1126/science.abi9009
    [14]
    Moradi M, Tajkhorshid E. Mechanistic picture for conformational transition of a membrane transporter at atomic resolution. Proc. Natl. Acad. Sci. U.S.A., 2013, 110 (47): 18916–18921. doi: 10.1073/pnas.1313202110
    [15]
    Wang Z, Liao J L. Probing structural determinants of ATP-binding cassette exporter conformational transition using coarse-grained molecular dynamics. J. Phys. Chem. B, 2015, 119 (4): 1295–1301. doi: 10.1021/jp509178k
    [16]
    Kieuvongngam V, Chen J. Structures of the peptidase-containing ABC transporter PCAT1 under equilibrium and nonequilibrium conditions. Proc. Natl. Acad. Sci. U.S.A., 2022, 119 (4): e2120534119. doi: 10.1073/pnas.2120534119
    [17]
    Abraham M J, Murtola T, Schulz R, et al. GROMACS: High performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX, 2015, 1–2: 19–25. doi: 10.1016/j.softx.2015.06.001
    [18]
    Simpson B W, Trent M S. Pushing the envelope: LPS modifications and their consequences. Nat. Rev. Microbiol., 2019, 17 (7): 403–416. doi: 10.1038/s41579-019-0201-x
    [19]
    Marrink S J, Risselada H J, Yefimov S, et al. The MARTINI force field: Coarse grained model for biomolecular simulations. J. Phys. Chem. B, 2007, 111 (27): 7812–7824. doi: 10.1021/jp071097f
    [20]
    Monticelli L, Kandasamy S K, Periole X, et al. The MARTINI coarse-grained force field: Extension to proteins. J. Chem. Theory Comput., 2008, 4 (5): 819–834. doi: 10.1021/ct700324x
    [21]
    López C A, Sovova Z, van Eerden F J, et al. Martini force field parameters for glycolipids. J. Chem. Theory Comput., 2013, 9 (3): 1694–1708. doi: 10.1021/ct3009655
    [22]
    Darden T, York D, Pedersen L. Particle mesh Ewald: An N·log(N) method for Ewald sums in large systems. J. Chem. Phys., 1993, 98: 10089–10092. doi: 10.1063/1.464397
    [23]
    Huang Y, Xu H C, Liao J L. Coarse-grained free-energy simulations of conformational state transitions in an adenosine 5′-triphosphate-binding cassette exporter. Chin. J. Chem. Phys., 2020, 33: 712–716. doi: 10.1063/1674-0068/cjcp1908149
    [24]
    Periole X, Knepp A M, Sakmar T P, et al. Structural determinants of the supramolecular organization of G protein-coupled receptors in bilayers. J. Am. Chem. Soc., 2012, 134 (26): 10959–10965. doi: 10.1021/ja303286e
    [25]
    Lemkul J A, Bevan D R. Assessing the stability of Alzheimer’s amyloid protofibrils using molecular dynamics. J. Phys. Chem. B, 2010, 114 (4): 1652–1660. doi: 10.1021/jp9110794
    [26]
    Jakubec D, Vondrášek J. Efficient estimation of absolute binding free energy for a homeodomain-DNA complex from nonequilibrium pulling simulations. J. Chem. Theory Comput., 2020, 16 (4): 2034–2041. doi: 10.1021/acs.jctc.0c00006
    [27]
    Xing X, Liu C, Ali A, et al. Novel disassembly mechanisms of sigmoid Aβ42 protofibrils by introduced neutral and charged drug molecules. ACS Chem. Neuro., 2020, 11 (1): 45–56. doi: 10.1021/acschemneuro.9b00550
    [28]
    Wassenaar T A, Pluhackova K, Böckmann R A, et al. Going backward: A flexible geometric approach to reverse transformation from coarse grained to atomistic models. J. Chem. Theory Comput., 2014, 10 (2): 676–690. doi: 10.1021/ct400617g
    [29]
    Hohl M, Briand C, Grütter M G, et al. Crystal structure of a heterodimeric ABC transporter in its inward-facing conformation. Nat. Struct. Mol. Biol., 2012, 19 (4): 395–402. doi: 10.1038/nsmb.2267
    [30]
    Jin M S, Oldham M L, Zhang Q, et al. Crystal structure of the multidrug transporter P-glycoprotein from Caenorhabditis elegans. Nature, 2012, 490 (7421): 566–569. doi: 10.1038/nature11448
    [31]
    Huang W, Liao J L. Catalytic mechanism of the maltose transporter hydrolyzing ATP. Biochemistry, 2016, 55 (1): 224–231. doi: 10.1021/acs.biochem.5b00970
    • Supplements(1)
  • JUSTC-2022-0134 Supporting information.docx
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    Figure  1.  Three conformations of an ABC exporter. (a) Inward-facing (IF) conformation, in which the internal gate is open whereas the external gate is closed. (b) Occluded (OC) conformation, in which both the internal and external gates are closed. (c) Outward-facing (OF) conformation, in which the internal gate is closed whereas the external gate is open. The TMD helices, TM1−TM6 on one subunit and TM1′−TM6′ on the other, are colored red and blue, respectively. The intracellular coupling helices ICH1 (ICH1′), which links TM2 (TM2′) and TM3 (TM3′) at its N- and C-terminus, and ICH2 (ICH2′), which links TM4 (TM4′) and TM5 (TM5′), are colored green and yellow, respectively.

    Figure  2.  Potential of mean force (PMF) expressed as a function of the COM distances, d1, for the internal gate, and d2, for the external gate. (a) d1=2.48 nm and d2=2.83 nm, (b) d1=2.61 nm and d2=2.42 nm, (c) d1=3.21 nm and d2=2.39 nm, and (d) d1=3.48–3.82 nm and d2=2.34 nm represent the OF, OC, IF1, and IF2 states, respectively. The coarse-grained structures, which represent the (a) OF, (b) OC, (c) IF1, and (d) IF2 states, and their corresponding atomistic structures are also presented.

    Figure  3.  Time evolution of d1, d2, dNBD, dICH1 and dICH2 upon NBD dissociation.

    Figure  4.  (A) Coarse-grained structures from the CG-MD simulations for the (a) OF, (b) OC, (c) IF1, and (d) IF2 states and (B) a mechanistic model for conformational state transitions in response to NBD dissociation. For clarity, two NBDs are presented with two balls (red and blue), and only TM3 and TM4-TM5 (red) and TM3′ and TM4′-TM5′ (blue) are presented with rectangular sticks. The COM distances d1 and d2 are displayed in (A). In (B), the wider rectangular sticks represent TM4-TM5 (red, TM2′ not shown) and TM4′-TM5′ (blue, TM2 not shown), whereas the narrower rectangular sticks represent TM3 (red) and TM3′ (blue). In (B), symbols $ \leftrightarrow $ and ←→ represent the gate opening, whereas →← represents the closing of a gate.

    [1]
    Thomas C, Tampé R. Structural and mechanistic principles of ABC transporters. Annu. Rev. Biochem., 2020, 89: 605–636. doi: 10.1146/annurev-biochem-011520-105201
    [2]
    Srikant S, Gaudet R. Mechanics and pharmacology of substrate selection and transport by eukaryotic ABC exporters. Nat. Struct. Mol. Biol., 2019, 26 (9): 792–801. doi: 10.1038/s41594-019-0280-4
    [3]
    Hofmann S, Januliene D, Mehdipour A R, et al. Conformation space of a heterodimeric ABC exporter under turnover conditions. Nature, 2019, 571 (7766): 580–583. doi: 10.1038/s41586-019-1391-0
    [4]
    Locher K P. Mechanistic diversity in ATP-binding cassette (ABC) transporters. Nat. Struct. Mol. Biol., 2016, 23 (6): 487–493. doi: 10.1038/nsmb.3216
    [5]
    Robey R W, Pluchino K M, Hall M D, et al. Revisiting the role of ABC transporters in multidrug-resistant cancer. Nat. Rev. Cancer, 2018, 18 (7): 452–464. doi: 10.1038/s41568-018-0005-8
    [6]
    Dawson R J P, Locher K P. Structure of a bacterial multidrug ABC transporter. Nature, 2006, 443 (7108): 180–185. doi: 10.1038/nature05155
    [7]
    Dawson R J P, Locher K P. Structure of the multidrug ABC transporter Sav1866 from Staphylococcus aureus in complex with AMP-PNP. FEBS Lett., 2007, 581 (5): 935–938. doi: 10.1016/j.febslet.2007.01.073
    [8]
    Ward A, Reyes C L, Yu J, et al. Flexibility in the ABC transporter MsbA: Alternating access with a twist. Proc. Natl. Acad. Sci. U.S.A., 2007, 104 (48): 19005–19010. doi: 10.1073/pnas.0709388104
    [9]
    Mi W, Li Y, Yoon S H, et al. Structural basis of MsbA-mediated lipopolysaccharide transport. Nature, 2017, 549 (7671): 233–237. doi: 10.1038/nature23649
    [10]
    Ho H, Miu A, Alexander M K, et al. Structural basis for dual-mode inhibition of the ABC transporter MsbA. Nature, 2018, 557 (7704): 196–201. doi: 10.1038/s41586-018-0083-5
    [11]
    Padayatti P S, Lee S C, Stanfield R L, et al. Structural insights into the lipid A transport pathway in MsbA. Structure, 2019, 27 (7): 1114–1123.e3. doi: 10.1016/j.str.2019.04.007
    [12]
    Angiulli G, Dhupar H S, Suzuki H, et al. New approach for membrane protein reconstitution into peptidiscs and basis for their adaptability to different proteins. eLife, 2020, 9: e53530. doi: 10.7554/eLife.53530
    [13]
    Thélot F A, Zhang W, Song K, et al. Distinct allosteric mechanisms of first-generation MsbA inhibitors. Science, 2021, 374 (6567): 580–585. doi: 10.1126/science.abi9009
    [14]
    Moradi M, Tajkhorshid E. Mechanistic picture for conformational transition of a membrane transporter at atomic resolution. Proc. Natl. Acad. Sci. U.S.A., 2013, 110 (47): 18916–18921. doi: 10.1073/pnas.1313202110
    [15]
    Wang Z, Liao J L. Probing structural determinants of ATP-binding cassette exporter conformational transition using coarse-grained molecular dynamics. J. Phys. Chem. B, 2015, 119 (4): 1295–1301. doi: 10.1021/jp509178k
    [16]
    Kieuvongngam V, Chen J. Structures of the peptidase-containing ABC transporter PCAT1 under equilibrium and nonequilibrium conditions. Proc. Natl. Acad. Sci. U.S.A., 2022, 119 (4): e2120534119. doi: 10.1073/pnas.2120534119
    [17]
    Abraham M J, Murtola T, Schulz R, et al. GROMACS: High performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX, 2015, 1–2: 19–25. doi: 10.1016/j.softx.2015.06.001
    [18]
    Simpson B W, Trent M S. Pushing the envelope: LPS modifications and their consequences. Nat. Rev. Microbiol., 2019, 17 (7): 403–416. doi: 10.1038/s41579-019-0201-x
    [19]
    Marrink S J, Risselada H J, Yefimov S, et al. The MARTINI force field: Coarse grained model for biomolecular simulations. J. Phys. Chem. B, 2007, 111 (27): 7812–7824. doi: 10.1021/jp071097f
    [20]
    Monticelli L, Kandasamy S K, Periole X, et al. The MARTINI coarse-grained force field: Extension to proteins. J. Chem. Theory Comput., 2008, 4 (5): 819–834. doi: 10.1021/ct700324x
    [21]
    López C A, Sovova Z, van Eerden F J, et al. Martini force field parameters for glycolipids. J. Chem. Theory Comput., 2013, 9 (3): 1694–1708. doi: 10.1021/ct3009655
    [22]
    Darden T, York D, Pedersen L. Particle mesh Ewald: An N·log(N) method for Ewald sums in large systems. J. Chem. Phys., 1993, 98: 10089–10092. doi: 10.1063/1.464397
    [23]
    Huang Y, Xu H C, Liao J L. Coarse-grained free-energy simulations of conformational state transitions in an adenosine 5′-triphosphate-binding cassette exporter. Chin. J. Chem. Phys., 2020, 33: 712–716. doi: 10.1063/1674-0068/cjcp1908149
    [24]
    Periole X, Knepp A M, Sakmar T P, et al. Structural determinants of the supramolecular organization of G protein-coupled receptors in bilayers. J. Am. Chem. Soc., 2012, 134 (26): 10959–10965. doi: 10.1021/ja303286e
    [25]
    Lemkul J A, Bevan D R. Assessing the stability of Alzheimer’s amyloid protofibrils using molecular dynamics. J. Phys. Chem. B, 2010, 114 (4): 1652–1660. doi: 10.1021/jp9110794
    [26]
    Jakubec D, Vondrášek J. Efficient estimation of absolute binding free energy for a homeodomain-DNA complex from nonequilibrium pulling simulations. J. Chem. Theory Comput., 2020, 16 (4): 2034–2041. doi: 10.1021/acs.jctc.0c00006
    [27]
    Xing X, Liu C, Ali A, et al. Novel disassembly mechanisms of sigmoid Aβ42 protofibrils by introduced neutral and charged drug molecules. ACS Chem. Neuro., 2020, 11 (1): 45–56. doi: 10.1021/acschemneuro.9b00550
    [28]
    Wassenaar T A, Pluhackova K, Böckmann R A, et al. Going backward: A flexible geometric approach to reverse transformation from coarse grained to atomistic models. J. Chem. Theory Comput., 2014, 10 (2): 676–690. doi: 10.1021/ct400617g
    [29]
    Hohl M, Briand C, Grütter M G, et al. Crystal structure of a heterodimeric ABC transporter in its inward-facing conformation. Nat. Struct. Mol. Biol., 2012, 19 (4): 395–402. doi: 10.1038/nsmb.2267
    [30]
    Jin M S, Oldham M L, Zhang Q, et al. Crystal structure of the multidrug transporter P-glycoprotein from Caenorhabditis elegans. Nature, 2012, 490 (7421): 566–569. doi: 10.1038/nature11448
    [31]
    Huang W, Liao J L. Catalytic mechanism of the maltose transporter hydrolyzing ATP. Biochemistry, 2016, 55 (1): 224–231. doi: 10.1021/acs.biochem.5b00970
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