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Flexible Gates Generate Occluded Intermediates in the Transport Cycle of LacY()

The major facilitator superfamily (MFS) transporter lactose permease (LacY) alternates between cytoplasmic and periplasmic open conformations to co-transport a sugar molecule together with a proton across the plasma membrane. Indirect experimental evidence suggested the existence of an occluded tran...

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Autores principales: Stelzl, Lukas S., Fowler, Philip W., Sansom, Mark S.P., Beckstein, Oliver
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3905165/
https://www.ncbi.nlm.nih.gov/pubmed/24513108
http://dx.doi.org/10.1016/j.jmb.2013.10.024
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author Stelzl, Lukas S.
Fowler, Philip W.
Sansom, Mark S.P.
Beckstein, Oliver
author_facet Stelzl, Lukas S.
Fowler, Philip W.
Sansom, Mark S.P.
Beckstein, Oliver
author_sort Stelzl, Lukas S.
collection PubMed
description The major facilitator superfamily (MFS) transporter lactose permease (LacY) alternates between cytoplasmic and periplasmic open conformations to co-transport a sugar molecule together with a proton across the plasma membrane. Indirect experimental evidence suggested the existence of an occluded transition intermediate of LacY, which would prevent leaking of the proton gradient. As no experimental structure is known, the conformational transition is not fully understood in atomic detail. We simulated transition events from a cytoplasmic open conformation to a periplasmic open conformation with the dynamic importance sampling molecular dynamics method and observed occluded intermediates. Analysis of water permeation pathways and the electrostatic free-energy landscape of a solvated proton indicated that the occluded state contains a solvated central cavity inaccessible from either side of the membrane. We propose a pair of geometric order parameters that capture the state of the pathway through the MFS transporters as shown by a survey of available crystal structures and models. We present a model for the occluded state of apo-LacY, which is similar to the occluded crystal structures of the MFS transporters EmrD, PepT(So), NarU, PiPT and XylE. Our simulations are consistent with experimental double electron spin–spin distance measurements that have been interpreted to show occluded conformations. During the simulations, a salt bridge that has been postulated to be involved in driving the conformational transition formed. Our results argue against a simple rigid-body domain motion as implied by a strict “rocker-switch mechanism” and instead hint at an intricate coupling between two flexible gates.
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spelling pubmed-39051652014-02-06 Flexible Gates Generate Occluded Intermediates in the Transport Cycle of LacY() Stelzl, Lukas S. Fowler, Philip W. Sansom, Mark S.P. Beckstein, Oliver J Mol Biol Article The major facilitator superfamily (MFS) transporter lactose permease (LacY) alternates between cytoplasmic and periplasmic open conformations to co-transport a sugar molecule together with a proton across the plasma membrane. Indirect experimental evidence suggested the existence of an occluded transition intermediate of LacY, which would prevent leaking of the proton gradient. As no experimental structure is known, the conformational transition is not fully understood in atomic detail. We simulated transition events from a cytoplasmic open conformation to a periplasmic open conformation with the dynamic importance sampling molecular dynamics method and observed occluded intermediates. Analysis of water permeation pathways and the electrostatic free-energy landscape of a solvated proton indicated that the occluded state contains a solvated central cavity inaccessible from either side of the membrane. We propose a pair of geometric order parameters that capture the state of the pathway through the MFS transporters as shown by a survey of available crystal structures and models. We present a model for the occluded state of apo-LacY, which is similar to the occluded crystal structures of the MFS transporters EmrD, PepT(So), NarU, PiPT and XylE. Our simulations are consistent with experimental double electron spin–spin distance measurements that have been interpreted to show occluded conformations. During the simulations, a salt bridge that has been postulated to be involved in driving the conformational transition formed. Our results argue against a simple rigid-body domain motion as implied by a strict “rocker-switch mechanism” and instead hint at an intricate coupling between two flexible gates. Elsevier 2014-02-06 /pmc/articles/PMC3905165/ /pubmed/24513108 http://dx.doi.org/10.1016/j.jmb.2013.10.024 Text en © 2013 The Authors https://creativecommons.org/licenses/by/3.0/This is an open access article under the CC BY license (https://creativecommons.org/licenses/by/3.0/).
spellingShingle Article
Stelzl, Lukas S.
Fowler, Philip W.
Sansom, Mark S.P.
Beckstein, Oliver
Flexible Gates Generate Occluded Intermediates in the Transport Cycle of LacY()
title Flexible Gates Generate Occluded Intermediates in the Transport Cycle of LacY()
title_full Flexible Gates Generate Occluded Intermediates in the Transport Cycle of LacY()
title_fullStr Flexible Gates Generate Occluded Intermediates in the Transport Cycle of LacY()
title_full_unstemmed Flexible Gates Generate Occluded Intermediates in the Transport Cycle of LacY()
title_short Flexible Gates Generate Occluded Intermediates in the Transport Cycle of LacY()
title_sort flexible gates generate occluded intermediates in the transport cycle of lacy()
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3905165/
https://www.ncbi.nlm.nih.gov/pubmed/24513108
http://dx.doi.org/10.1016/j.jmb.2013.10.024
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