Sequence and structural determinants of ligand-dependent alternating access of a MATE transporter.
Proc Natl Acad Sci U S A
; 117(9): 4732-4740, 2020 03 03.
Article
em En
| MEDLINE
| ID: mdl-32075917
ABSTRACT
Multidrug and toxic compound extrusion (MATE) transporters are ubiquitous ion-coupled antiporters that extrude structurally and chemically dissimilar cytotoxic compounds and have been implicated in conferring multidrug resistance. Here, we integrate double electron-electron resonance (DEER) with functional assays and site-directed mutagenesis of conserved residues to illuminate principles of ligand-dependent alternating access of PfMATE, a proton-coupled MATE from the hyperthermophilic archaeon Pyrococcus furiosus Pairs of spin labels monitoring the two sides of the transporter reconstituted into nanodiscs reveal large-amplitude movement of helices that alter the orientation of a putative substrate binding cavity. We found that acidic pH favors formation of an inward-facing (IF) conformation, whereas elevated pH (>7) and the substrate rhodamine 6G stabilizes an outward-facing (OF) conformation. The lipid-dependent PfMATE isomerization between OF and IF conformation is driven by protonation of a previously unidentified intracellular glutamate residue that is critical for drug resistance. Our results can be framed in a mechanistic model of transport that addresses central aspects of ligand coupling and alternating access.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Antiporters
/
Proteínas de Transporte de Cátions Orgânicos
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article