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Mode of Interaction of the Signal-Transducing Protein EIIAGlc with the Maltose ABC Transporter in the Process of Inducer Exclusion

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posted on 29.08.2016 by Steven Wuttge, Anke Licht, M. Hadi Timachi, Enrica Bordignon, Erwin Schneider
Enzyme IIAGlc (EIIAGlc) of the phosphoenolpyruvate phosphotransferase system for the uptake of glucose in Escherichia coli and Salmonella inhibits the maltose ATP-binding cassette transporter (MalE-FGK2) by interaction with the nucleotide-binding and -hydrolyzing subunit MalK, a process termed inducer exclusion. We have investigated binding of EIIAGlc to the MalK dimer by cysteine cross-linking in proteoliposomes. The results prove that the binding site I of EIIAGlc is contacting the N-terminal subdomain of MalK while the binding site II is relatively close to the C-terminal (regulatory) subdomain, in agreement with a crystal structure [Chen, S., Oldham, M. L., Davidson, A. L., and Chen, J. (2013) Nature 499, 364−368]. Moreover, EIIAGlc was found to bind to the MalK dimer regardless of its conformational state. Deletion of the amphipathic N-terminal peptide of EIIAGlc, which is required for inhibition, reduced formation of cross-linked products. Using a spin-labeled transporter variant and EPR spectroscopy, we demonstrate that EIIAGlc arrests the transport cycle by inhibiting the ATP-dependent closure of the MalK dimer.

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