Dataset Information

Flexible-to-rigid transition is central for substrate transport in the ABC transporter BmrA from Bacillus subtilis.

ABSTRACT: ATP-binding-cassette (ABC) transporters are molecular pumps that translocate molecules across the cell membrane by switching between inward-facing and outward-facing states. To obtain a detailed understanding of their mechanism remains a challenge to structural biology, as these proteins are notoriously difficult to study at the molecular level in their active, membrane-inserted form. Here we use solid-state NMR to investigate the multidrug ABC transporter BmrA reconstituted in lipids. We identify the chemical-shift differences between the inward-facing, and outward-facing state induced by ATP:Mg2+:Vi addition. Analysis of an X-loop mutant, for which we show that ATPase and transport activities are uncoupled, reveals an incomplete transition to the outward-facing state upon ATP:Mg2+:Vi addition, notably lacking the decrease in dynamics of a defined set of residues observed in wild-type BmrA. This suggests that this stiffening is required for an efficient transmission of the conformational changes to allow proper transport of substrate by the pump.

SUBMITTER: Lacabanne D

PROVIDER: S-EPMC6488656 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Flexible-to-rigid transition is central for substrate transport in the ABC transporter BmrA from <i>Bacillus subtilis</i>.

Lacabanne Denis D Orelle Cédric C Lecoq Lauriane L Kunert Britta B Chuilon Claire C Wiegand Thomas T Ravaud Stéphanie S Jault Jean-Michel JM Meier Beat H BH Böckmann Anja A

Communications biology 20190429

ATP-binding-cassette (ABC) transporters are molecular pumps that translocate molecules across the cell membrane by switching between inward-facing and outward-facing states. To obtain a detailed understanding of their mechanism remains a challenge to structural biology, as these proteins are notoriously difficult to study at the molecular level in their active, membrane-inserted form. Here we use solid-state NMR to investigate the multidrug ABC transporter BmrA reconstituted in lipids. We identi ...[more]

PMID: 31044174

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Project description:The accumulation of compatible solutes is a common defense of bacteria against the detrimental effects of high osmolarity. Uptake systems for these compounds are cornerstones in cellular osmostress responses because they allow the energy-preserving scavenging of osmostress protectants from environmental sources. Bacillus subtilis is well studied with respect to the import of compatible solutes and its five transport systems (OpuA, OpuB, OpuC, OpuD, and OpuE), for these stress protectants have previously been comprehensively studied. Building on this knowledge and taking advantage of the unabated appearance of new genome sequences of members of the genus Bacillus, we report here the discovery, physiological characterization, and phylogenomics of a new member of the Opu family of transporters, OpuF (OpuFA-OpuFB). OpuF is not present in B. subtilis but it is widely distributed in members of the large genus Bacillus OpuF is a representative of a subgroup of ATP-binding cassette (ABC) transporters in which the substrate-binding protein (SBP) is fused to the transmembrane domain (TMD). We studied the salient features of the OpuF transporters from Bacillus infantis and Bacillus panaciterrae by functional reconstitution in a B. subtilis chassis strain lacking known Opu transporters. A common property of the examined OpuF systems is their substrate profile; OpuF mediates the import of glycine betaine, proline betaine, homobetaine, and the marine osmolyte dimethylsulfoniopropionate (DMSP). An in silico model of the SBP domain of the TMD-SBP hybrid protein OpuFB was established. It revealed the presence of an aromatic cage, a structural feature commonly present in ligand-binding sites of compatible solute importers.IMPORTANCE The high-affinity import of compatible solutes from environmental sources is an important aspect of the cellular defense of many bacteria and archaea against the harmful effects of high external osmolarity. The accumulation of these osmostress protectants counteracts high-osmolarity-instigated water efflux, a drop in turgor to nonphysiological values, and an undue increase in molecular crowding of the cytoplasm; they thereby foster microbial growth under osmotically unfavorable conditions. Importers for compatible solutes allow the energy-preserving scavenging of osmoprotective and physiologically compliant organic solutes from environmental sources. We report here the discovery, exemplary physiological characterization, and phylogenomics of a new compatible solute importer, OpuF, widely found in members of the Bacillus genus. The OpuF system is a representative of a growing subgroup of ABC transporters in which the substrate-scavenging function of the substrate-binding protein (SBP) and the membrane-embedded substrate translocating subunit (TMD) are fused into a single polypeptide chain.

Dataset Information

Flexible-to-rigid transition is central for substrate transport in the ABC transporter BmrA from Bacillus subtilis.

Publications

Flexible-to-rigid transition is central for substrate transport in the ABC transporter BmrA from <i>Bacillus subtilis</i>.

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