Both maltose-binding protein and ATP are required for nucleotide-binding domain closure in the intact maltose ABC transporter

The maltose transporter MalFGK₂ of Escherichia coli is a member of the ATP-binding cassette superfamily. A periplasmic maltose-binding protein (MBP) delivers maltose to MalFGK₂ 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 MalFGK₂ 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 Pi release coincides with the reorientation of transmembrane helices to an inward-facing conformation and the final step of maltose translocation into the cell.