Sugar transport by the bacterial phosphotransferase system. Characterization of the Escherichia coli enzyme I monomer/dimer transition kinetics by fluorescence anisotropy.

Enzyme I of the bacterial phosphoenolpyruvate: glycose phosphotransferase system (PTS) exists in a monomer/dimer (M/D) equilibrium. These two forms are functionally different, and their interconversion may be a means of regulating the PTS. The M/D equilibrium was studied by fluorescence anisotropy of a pyrene derivative (Chauvin, F., Brand, L., and Roseman, S. (1994) J. Biol. Chem. 269, 20263-20269). In this paper, the kinetics of the transition is investigated. The following apparent rate constants were found for the M/D transition of phospho-Enzyme I in the presence of Mg2+ and PEP at 6 degrees C: kA = 3.4 x 10(3) M-1 s-1 and kD = 1.04 x 10(3) s-1. The association rate is especially slow, 2-3 orders of magnitude slower than the average dimerization rate determined for other proteins. Furthermore, the rate of quaternary structure changes matches that of enzymatic activity changes, as well as that of tertiary structure changes (Chauvin, F., Toptygin, D., Roseman, S., and Brand, L. (1992) Biophys. Chem. 44, 163-173). Finally, the effect of two ligands is shown; PEP increases the relaxation rate by 3-fold at 23 degrees C, and Mg2+ addition causes a 4-fold increase in the relaxation rate.