Solution Structure of the Phosphoryl Transfer Complex between the Signal-transducing Protein IIAGlucose and the Cytoplasmic Domain of the Glucose Transporter IICBGlucose of the Escherichia coli Glucose Phosphotransferase System

Mengli Cai, David C. Williams, Guangshun Wang, Byeong Ryong Lee, Alan Peterkofsky, G. Marius Clore

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59 Scopus citations

Abstract

The solution structure of the final phosphoryl transfer complex in the glucose-specific arm of the Escherichia coli phosphotransferase system, between enzyme IIAGlucose (IIAGlc) and the cytoplasmic B domain (IIBGlc) of the glucose transporter IICBGlc, has been solved by NMR. The interface (∼1200-Å2 buried surface) is formed by the interaction of a concave depression on IIAGlc with a convex protrusion on IIBGlc. The phosphoryl donor and acceptor residues, His-90 of IIAGlc and Cys-35 of IIBGlc (residues of IIBGlc are denoted in italics) are in close proximity and buried at the center of the interface. Cys-35 is primed for nucleophilic attack on the phosphorus atom by stabilization of the thiolate anion (pKa ∼6.5) through intramolecular hydrogen bonding interactions with several adjacent backbone amide groups. Hydrophobic intermolecular contacts are supplemented by peripheral electrostatic interactions involving an alternating distribution of positively and negatively charged residues on the interaction surfaces of both proteins. Salt bridges between the Asp-38/Asp-94 pair of IIAGlc and the Arg-38/Arg-40 pair of IIBGlc neutralize the accumulation of negative charge in the vicinity of both the Sγ atom of Cys-35 and the phosphoryl group in the complex. A pentacoordinate phosphoryl transition state is readily accommodated without any change in backbone conformation, and the structure of the complex accounts for the preferred directionality of phosphoryl transfer between IIAGlc and IIBGlc. The structures of IIAGlc·IIBGlc and the two upstream complexes of the glucose phosphotransferase system (EI·HPr and IIA Glc·HPr) reveal a cascade in which highly overlapping binding sites on HPr and IIAGlc recognize structurally diverse proteins.

Original languageEnglish (US)
Pages (from-to)25191-25206
Number of pages16
JournalJournal of Biological Chemistry
Volume278
Issue number27
DOIs
StatePublished - Jul 4 2003

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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