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Residues forming the gating regions of asymmetric multidrug transporter Pdr5 also play roles in conformational switching and protein folding

ATP-binding cassette (ABC) multidrug transporters are large, polytopic membrane proteins that exhibit astonishing promiscuity for their transport substrates. These transporters unidirectionally efflux thousands of structurally and functionally distinct compounds. To preclude the reentry of xenobioti...

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Autores principales: Alhumaidi, Maryam, Nentwig, Lea-Marie, Rahman, Hadiar, Schmitt, Lutz, Rudrow, Andrew, Harris, Andrzej, Dillon, Cierra, Restrepo, Lucas, Lamping, Erwin, Arya, Nidhi, Ambudkar, Suresh V., Choy, John S., Golin, John
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society for Biochemistry and Molecular Biology 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9723933/
https://www.ncbi.nlm.nih.gov/pubmed/36370844
http://dx.doi.org/10.1016/j.jbc.2022.102689
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author Alhumaidi, Maryam
Nentwig, Lea-Marie
Rahman, Hadiar
Schmitt, Lutz
Rudrow, Andrew
Harris, Andrzej
Dillon, Cierra
Restrepo, Lucas
Lamping, Erwin
Arya, Nidhi
Ambudkar, Suresh V.
Choy, John S.
Golin, John
author_facet Alhumaidi, Maryam
Nentwig, Lea-Marie
Rahman, Hadiar
Schmitt, Lutz
Rudrow, Andrew
Harris, Andrzej
Dillon, Cierra
Restrepo, Lucas
Lamping, Erwin
Arya, Nidhi
Ambudkar, Suresh V.
Choy, John S.
Golin, John
author_sort Alhumaidi, Maryam
collection PubMed
description ATP-binding cassette (ABC) multidrug transporters are large, polytopic membrane proteins that exhibit astonishing promiscuity for their transport substrates. These transporters unidirectionally efflux thousands of structurally and functionally distinct compounds. To preclude the reentry of xenobiotic molecules via the drug-binding pocket, these proteins contain a highly conserved molecular gate, essentially allowing the transporters to function as molecular diodes. However, the structure–function relationship of these conserved gates and gating regions are not well characterized. In this study, we combine recent single-molecule, cryo-EM data with genetic and biochemical analyses of residues in the gating region of the yeast multidrug transporter Pdr5, the founding member of a large group of clinically relevant asymmetric ABC efflux pumps. Unlike the symmetric ABCG2 efflux gate, the Pdr5 counterpart is highly asymmetric, with only four (instead of six) residues comprising the gate proper. However, other residues in the near vicinity are essential for the gating activity. Furthermore, we demonstrate that residues in the gate and in the gating regions have multiple functions. For example, we show that Ile-685 and Val-1372 are required not only for successful efflux but also for allosteric inhibition of Pdr5 ATPase activity. Our investigations reveal that the gating region residues of Pdr5, and possibly other ABCG transporters, play a role not only in molecular gating but also in allosteric regulation, conformational switching, and protein folding.
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spelling pubmed-97239332022-12-07 Residues forming the gating regions of asymmetric multidrug transporter Pdr5 also play roles in conformational switching and protein folding Alhumaidi, Maryam Nentwig, Lea-Marie Rahman, Hadiar Schmitt, Lutz Rudrow, Andrew Harris, Andrzej Dillon, Cierra Restrepo, Lucas Lamping, Erwin Arya, Nidhi Ambudkar, Suresh V. Choy, John S. Golin, John J Biol Chem Research Article ATP-binding cassette (ABC) multidrug transporters are large, polytopic membrane proteins that exhibit astonishing promiscuity for their transport substrates. These transporters unidirectionally efflux thousands of structurally and functionally distinct compounds. To preclude the reentry of xenobiotic molecules via the drug-binding pocket, these proteins contain a highly conserved molecular gate, essentially allowing the transporters to function as molecular diodes. However, the structure–function relationship of these conserved gates and gating regions are not well characterized. In this study, we combine recent single-molecule, cryo-EM data with genetic and biochemical analyses of residues in the gating region of the yeast multidrug transporter Pdr5, the founding member of a large group of clinically relevant asymmetric ABC efflux pumps. Unlike the symmetric ABCG2 efflux gate, the Pdr5 counterpart is highly asymmetric, with only four (instead of six) residues comprising the gate proper. However, other residues in the near vicinity are essential for the gating activity. Furthermore, we demonstrate that residues in the gate and in the gating regions have multiple functions. For example, we show that Ile-685 and Val-1372 are required not only for successful efflux but also for allosteric inhibition of Pdr5 ATPase activity. Our investigations reveal that the gating region residues of Pdr5, and possibly other ABCG transporters, play a role not only in molecular gating but also in allosteric regulation, conformational switching, and protein folding. American Society for Biochemistry and Molecular Biology 2022-11-10 /pmc/articles/PMC9723933/ /pubmed/36370844 http://dx.doi.org/10.1016/j.jbc.2022.102689 Text en © 2022 The Authors https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Research Article
Alhumaidi, Maryam
Nentwig, Lea-Marie
Rahman, Hadiar
Schmitt, Lutz
Rudrow, Andrew
Harris, Andrzej
Dillon, Cierra
Restrepo, Lucas
Lamping, Erwin
Arya, Nidhi
Ambudkar, Suresh V.
Choy, John S.
Golin, John
Residues forming the gating regions of asymmetric multidrug transporter Pdr5 also play roles in conformational switching and protein folding
title Residues forming the gating regions of asymmetric multidrug transporter Pdr5 also play roles in conformational switching and protein folding
title_full Residues forming the gating regions of asymmetric multidrug transporter Pdr5 also play roles in conformational switching and protein folding
title_fullStr Residues forming the gating regions of asymmetric multidrug transporter Pdr5 also play roles in conformational switching and protein folding
title_full_unstemmed Residues forming the gating regions of asymmetric multidrug transporter Pdr5 also play roles in conformational switching and protein folding
title_short Residues forming the gating regions of asymmetric multidrug transporter Pdr5 also play roles in conformational switching and protein folding
title_sort residues forming the gating regions of asymmetric multidrug transporter pdr5 also play roles in conformational switching and protein folding
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9723933/
https://www.ncbi.nlm.nih.gov/pubmed/36370844
http://dx.doi.org/10.1016/j.jbc.2022.102689
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