Solution structure of NPr,a bacterial signal-transducing protein that controls the phosphorylation state of the potassium transporter-regulating protein IIA~(Ntr)

摘要

A nitrogen-related signal transduction pathway,consisting of the three phosphotransfer proteins EI~(Ntr),NPr,and IIA~(Ntr),was discovered recently to regulate the uptake of K~+ in Escherichia coli.In particular,dephosphorylated IIA~(Ntr)inhibits the activity of the K~+ transporter TrkA.Since the phosphorylation state of IIA~(Ntr)is partially determined by its reversible phosphorylation by NPr,we have determined the three-dimensional structure of NPr by solution NMR spectroscopy.In total,we obtained 973 NOE-derived distance restraints,112 chemical shift-derived backbone angle restraints,and 35 hydrogen-bond restraints derived from temperature coefficients(wave).We propose that temperature wave is useful for identifying exposed beta-strands and assists in establishing protein folds based on chemical shifts.The deduced structure of NPr contains three alpha-helices and four beta-strands with the three helices all packed on the same face of the beta-sheet.The active site residue His 16 of NPr for phosphoryl transfer was found to be neutral and in the Ne2-H tautomeric state.There appears to be increased motion in the active site region of NPr compared to HPr,a homologous protein involved in the uptake and regulation of carbohydrate utilization.