Subunit structure and activity of the mannitol-specific enzyme II of the Escherichia coli phosphoenolpyruvate-dependent phosphotransferase system solubilized in detergent.
The original proposal of Saier stating that P-enolpyruvate-dependent mannitol phosphorylation is catalyzed by the monomeric form of the bacterial phosphotransferase enzyme IImtl, which would be the form predominantly existing in the phospholipid bilayer, whereas mannitol/mannitol-P exchange would depend on the transient formation of functional dimers, is refuted [Saier, M.H. (1980) J. Supramol. Struct. 14, 281-294]. The correct interpretation of the proportional relation between the rate of mannitol phosphorylation in the overall reaction and the enzyme concentration is that enzyme IImtl is dimeric under the conditions employed. Differences measured in the enzyme concentration dependency of the overall and exchange reactions were caused by different assay conditions. The dimer is favored over the monomer at high ionic strength and basic pH. Mg2+ ions bind specifically to enzyme IImtl, inducing dimerization. A complex formed by mixing inorganic phosphate, F-, and Mg2+ at sufficiently high concentrations inhibits enzyme IImtl, in part, by dissociation of the dimer. Enzyme IImtl was dimeric in 25 mM Tris, pH 7.6, and 5 mM Mg2+ over a large enzyme concentration range and under many different turnover conditions. The association/dissociation equilibrium was demonstrated in phosphate bufers, pH 6.3. The dimer was the most active form both in the overall and in the exchange reaction under the conditions assayed. The monomer was virtually inactive in mannitol/mannitol-P exchange but retained 25% of the activity in the overall reaction.
Lolkema
J S
JS
Department of Physical Chemistry, University of Groningen, The Netherlands.
Robillard
G T
GT
D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism*; D003902:Detergents; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D008274:Magnesium / Q000378:metabolism; D008353:Mannitol / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D015394:Molecular Structure; D009004:Monosaccharide Transport Proteins; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D003902:Detergents; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins; D008353:Mannitol; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C022358:mannitol PTS permease, E coli; D008274:Magnesium