Fig. 5.

Time course of selected ¹H_N/¹⁵N cross-peaks in ¹H-¹⁵N HSQC spectra of wild-type IIBA^Mtl during active intramolecular phosphoryl transfer subsequent to the depletion of PEP. (a) Ala-383; (b) Asp-432; (c) Gly-555; (d) Thr-556. The spectrum at 0 h, taken at the point of complete depletion of PEP, reflects the biphosphorylated state in which both the A and B domains are fully phosphorylated; the spectrum at 4 h represents fully dephosphorylated IIBA^Mtl. The effects of chemical exchange arising from reversible phosphoryl transfer between the A and B domains in species that are monophosphorylated are apparent in the spectra taken at intermediate times. The Δω_HN values were obtained from the chemical-shift differences between the spectra taken at 0 and 4 h (¹H frequency of 800 MHz).