Sequential Action of MalE and Maltose Allows Coupling ATP Hydrolysis to Translocation in the MalFGK2 Transporter

Bao, Huan; Dalal, Kush; Cytrynbaum, Eric N.; Duong, Franck

doi:10.1074/jbc.m115.671826

Cited by 18 publications

(22 citation statements)

References 50 publications

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“…6B), which is in accordance with our previous report that ATP cleavage is stimulated by MalE (5,23). Maltose further increases the rate of ATP hydrolysis by increasing the rate of ADP and P i release from the nucleotide binding pockets (23) (Fig. 6C).…”

Section: Discussionsupporting

confidence: 92%

Negative Stain Single-particle EM of the Maltose Transporter in Nanodiscs Reveals Asymmetric Closure of MalK2 and Catalytic Roles of ATP, MalE, and Maltose

Fabre

Bao

Innes

et al. 2017

Journal of Biological Chemistry

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The MalE-MalFGK complex is one of the best characterized members of the large and ubiquitous family of ATP-binding cassette (ABC) transporters. It is composed of a membrane-spanning heterodimer, MalF-MalG; a homodimeric ATPase, MalK; and a periplasmic maltose receptor, MalE. Opening and closure of MalK is coupled to conformational changes in MalF-MalG and the alternate exposition of the substrate-binding site to either side of the membrane. To further define this alternate access mechanism and the impact of ATP, MalE, and maltose on the conformation of the transporter during the transport cycle, we have reconstituted MalFGK in nanodiscs and analyzed its conformations under 10 different biochemical conditions using negative stain single-particle EM. EM map results (at 15-25 Å resolution) indicate that binding of ATP to MalK promotes an asymmetric, semi-closed conformation in accordance with the low ATPase activity of MalFGK In the presence of MalE, the MalK dimer becomes fully closed, gaining the ability to hydrolyze ATP. In the presence of ADP or maltose, MalE·MalFGK remains essentially in a semi-closed symmetric conformation, indicating that release of these ligands is required for the return to the initial state. Taken together, this structural information provides a rationale for the stimulation of MalK ATPase activity by MalE as well as by maltose.

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Section: Discussionsupporting

confidence: 92%

Negative Stain Single-particle EM of the Maltose Transporter in Nanodiscs Reveals Asymmetric Closure of MalK2 and Catalytic Roles of ATP, MalE, and Maltose

Fabre

Bao

Innes

et al. 2017

Journal of Biological Chemistry

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“…We speculate that this is a mechanism to effectively capture and transport rare essential substrates such as some B vitamins. Type I ABC importers for highly abundant nutrients, such as the maltose transporter MalFGK, seem to show tight coupling between ATP hydrolysis and substrate transport 21 . Possibly there is also no evolutionary drive to optimise transporters for rare substrates as total ATP consumption for these transporters is negligible compared to total cellular ATP consumption.…”

Section: Discussionmentioning

confidence: 99%

“…Is the substrate immediately transported, and is ATP merely required to reset the transporter 20 ? Why is the ATPase activity at least one order of magnitude higher compared to transport of vitamin B 12 —in other words, why is there no strong coupling between ATP hydrolysis and substrate translocation such as observed for the well-studied type I maltose importer MalFGK 21 ? In order to address these questions, we have employed single-molecule fluorescence techniques to follow individual BtuCD-F proteins reconstituted in liposomes through time and to directly observe steps of the transport cycle.…”

Section: Introductionmentioning

confidence: 99%

Single-molecule visualization of conformational changes and substrate transport in the vitamin B12 ABC importer BtuCD-F

2017

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ATP-binding cassette (ABC) transporters form the largest class of active membrane transport proteins. Binding and hydrolysis of ATP by their highly conserved nucleotide-binding domains drive conformational changes of the complex that mediate transport of substrate across the membrane. The vitamin B12 importer BtuCD-F in Escherichia coli is an extensively studied model system. The periplasmic soluble binding protein BtuF binds the ligand; the transmembrane and ATPase domains BtuCD mediate translocation. Here we report the direct observation at the single-molecule level of ATP, vitamin B12 and BtuF-induced events in the transporter complex embedded in liposomes. Single-molecule fluorescence imaging techniques reveal that membrane-embedded BtuCD forms a stable complex with BtuF, regardless of the presence of ATP and vitamin B12. We observe that a vitamin B12 molecule remains bound to the complex for tens of seconds, during which several ATP hydrolysis cycles can take place, before it is being transported across the membrane.

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“…The binding of unliganded MalE to the transporter increases ATP hydrolysis and P i release becomes rate limiting. Liganded MalE, however, stimulates release of P i [118].…”

Section: Step 3 and 4: Substrate Delivery And Import Mechanismmentioning

confidence: 91%

An integrated transport mechanism of the maltose ABC importer

Mächtel

Narducci

Griffith

et al. 2019

Research in Microbiology

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ATP-binding cassette (ABC) transporters use the energy of ATP hydrolysis to transport a large diversity of molecules actively across biological membranes. A combination of biochemical, biophysical, and structural studies has established the maltose transporter MalFGK2 as one of the best characterized proteins of the ABC family. MalF and MalG are the transmembrane domains, and two MalKs form a homodimer of nucleotide-binding domains. A periplasmic maltose-binding protein (MalE) delivers maltose and other maltodextrins to the transporter, and triggers its ATPase activity. Substrate import occurs in a unidirectional manner by ATP-driven conformational changes in MalK2 that allow alternating access of the substrate-binding site in MalF to each side of the membrane. In this review, we present an integrated molecular mechanism of the transport process considering all currently available information. Furthermore, we summarize remaining inconsistencies and outline possible future routes to decipher the full mechanistic details of transport by MalEFGK2 complex and that of related importer systems.

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Sequential Action of MalE and Maltose Allows Coupling ATP Hydrolysis to Translocation in the MalFGK2 Transporter

Cited by 18 publications

References 50 publications

Negative Stain Single-particle EM of the Maltose Transporter in Nanodiscs Reveals Asymmetric Closure of MalK2 and Catalytic Roles of ATP, MalE, and Maltose

Negative Stain Single-particle EM of the Maltose Transporter in Nanodiscs Reveals Asymmetric Closure of MalK2 and Catalytic Roles of ATP, MalE, and Maltose

Single-molecule visualization of conformational changes and substrate transport in the vitamin B12 ABC importer BtuCD-F

An integrated transport mechanism of the maltose ABC importer

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