TY - JOUR
T1 - A novel membrane anchor function for the N-terminal amphipathic sequence of the signal-transducing protein IIAGlucose of the Escherichia coli phosphotransferase system
AU - Wang, Guangshun
AU - Peterkofsky, Alan
AU - Clore, G. Marius
PY - 2000/12/22
Y1 - 2000/12/22
N2 - Enzyme IIAGlucose (IIAGlc) is a signal-transducing protein in the phosphotransferase system of Escherichia coli. Structural studies of free IIAGlc and the HPr-IIAGlc complex have shown that IIAGlc comprises a globular β-sheet sandwich core (residues 19-168) and a disordered N-terminal tail (residues 1-18). Although the presence of the N-terminal tail is not required for IIAGlc to accept a phosphorus from the histidine phosphocarrier protein HPr, its presence is essential for effective phosphotransfer from IIAGlc to the membrane-bound IIBC Glc. The sequence of the N-terminal tail suggests that it has the potential to form an amphipathic helix. Using CD, we demonstrate that a peptide, corresponding to the N-terminal 18 residues of IIAGlc, adopts a helical conformation in the presence of either the anionic lipid phosphatidylglycerol or a mixture of anionic E. coli lipids phosphatidylglycerol (25%) and phosphatidylethanolamine (75%). The peptide, however, is in a random coil state in the presence of the zwitterionic lipid phosphatidylcholine, indicating that electrostatic interactions play a role in the binding of the lipid to the peptide. In addition, we show that intact IIAGlc also interacts with anionic lipids, resulting in an increase in helicity, which can be directly attributed to the N-terminal segment. From these data we propose that IIAGlc comprises two functional domains: a folded domain containing the active site and capable of weakly interacting with the peripheral IIB domain of the membrane protein IIBCGlc; and the N-terminal tail, which interacts with the negatively charged E. coli membrane, thereby stabilizing the complex of IIAGlc with IIBCGlc. This stabilization is essential for the final step of the phosphoryl transfer cascade in the glucose transport pathway.
AB - Enzyme IIAGlucose (IIAGlc) is a signal-transducing protein in the phosphotransferase system of Escherichia coli. Structural studies of free IIAGlc and the HPr-IIAGlc complex have shown that IIAGlc comprises a globular β-sheet sandwich core (residues 19-168) and a disordered N-terminal tail (residues 1-18). Although the presence of the N-terminal tail is not required for IIAGlc to accept a phosphorus from the histidine phosphocarrier protein HPr, its presence is essential for effective phosphotransfer from IIAGlc to the membrane-bound IIBC Glc. The sequence of the N-terminal tail suggests that it has the potential to form an amphipathic helix. Using CD, we demonstrate that a peptide, corresponding to the N-terminal 18 residues of IIAGlc, adopts a helical conformation in the presence of either the anionic lipid phosphatidylglycerol or a mixture of anionic E. coli lipids phosphatidylglycerol (25%) and phosphatidylethanolamine (75%). The peptide, however, is in a random coil state in the presence of the zwitterionic lipid phosphatidylcholine, indicating that electrostatic interactions play a role in the binding of the lipid to the peptide. In addition, we show that intact IIAGlc also interacts with anionic lipids, resulting in an increase in helicity, which can be directly attributed to the N-terminal segment. From these data we propose that IIAGlc comprises two functional domains: a folded domain containing the active site and capable of weakly interacting with the peripheral IIB domain of the membrane protein IIBCGlc; and the N-terminal tail, which interacts with the negatively charged E. coli membrane, thereby stabilizing the complex of IIAGlc with IIBCGlc. This stabilization is essential for the final step of the phosphoryl transfer cascade in the glucose transport pathway.
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U2 - 10.1074/jbc.C000709200
DO - 10.1074/jbc.C000709200
M3 - Article
C2 - 11044440
AN - SCOPUS:0034704122
VL - 275
SP - 39811
EP - 39814
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 51
ER -