Abstract
The maltose transporter MalFGK(2) of Escherichia coli is a member of the ATP-binding cassette superfamily. A periplasmic maltose-binding protein (MBP) delivers maltose to MalFGK(2) and stimulates its ATPase activity. Site-directed spin labeling EPR spectroscopy was used to study the opening and closing of the nucleotide-binding interface of MalFGK(2) during the catalytic cycle. In the intact transporter, closure of the interface coincides not just with the binding of ATP, as seen with isolated nucleotide-binding domains, but requires both MBP and ATP, implying that MBP stimulates ATPase activity by promoting the closure of the nucleotide-binding interface. After ATP hydrolysis, with MgADP and MBP bound, the nucleotide-binding interface resides in a semi-open configuration distinct from the fully open configuration seen in the absence of any ligand. We propose that P(i) release coincides with the reorientation of transmembrane helices to an inward-facing conformation and the final step of maltose translocation into the cell.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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ATP-Binding Cassette Transporters / chemistry*
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ATP-Binding Cassette Transporters / metabolism*
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Adenosine Triphosphatases / metabolism
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Adenosine Triphosphate / metabolism*
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Adenosine Triphosphate / pharmacology
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Adenylyl Imidodiphosphate / pharmacology
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Binding Sites
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Carrier Proteins / metabolism*
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Carrier Proteins / pharmacology
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Catalysis / drug effects
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Dimerization
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Electron Spin Resonance Spectroscopy
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Escherichia coli / metabolism*
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Escherichia coli Proteins / chemistry*
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Escherichia coli Proteins / metabolism*
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Ligands
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Liposomes / metabolism
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Maltose / metabolism
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Maltose-Binding Proteins
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Models, Molecular
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Monosaccharide Transport Proteins / chemistry*
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Monosaccharide Transport Proteins / metabolism*
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Mutant Proteins / metabolism
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Protein Structure, Tertiary
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Spin Labels
Substances
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ATP-Binding Cassette Transporters
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Carrier Proteins
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Escherichia coli Proteins
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Ligands
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Liposomes
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Maltose-Binding Proteins
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Monosaccharide Transport Proteins
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Mutant Proteins
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Spin Labels
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maltose transport system, E coli
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Adenylyl Imidodiphosphate
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Maltose
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Adenosine Triphosphate
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Adenosine Triphosphatases