HPr(His approximately P)-mediated phosphorylation differently affects counterflow and proton motive force-driven uptake via the lactose transport protein of Streptococcus thermophilus.
The lactose transport protein (LacS) of Streptococcus thermophilus has a C-terminal hydrophilic domain that is homologous to IIA protein and protein domains of the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS). The IIA domain of LacS is phosphorylated on His-552 by the general energy coupling proteins of the PTS, which are Enzyme I and HPr. To study the effect of phosphorylation on transport, the LacS protein was purified and incorporated into liposomes with the IIA domain facing outwards. This allowed the phosphorylation of the membrane-reconstituted protein by purified HPr(His approximately P) of S. thermophilus. Phosphorylation of LacS increased the V(max) of counterflow transport, whereas the V(max) of the proton motive force (delta p)-driven lactose uptake was not affected. In line with a range of kinetic studies, we propose that phosphorylation affects the rate constants for the reorientation of the ternary complex (LacS with bound lactose plus proton), which is rate-determining for counterflow but not for delta p-driven transport.
Gunnewijk
M G
MG
Department of Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.
Poolman
B
B
D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000378:metabolism*; D010766:Phosphorylation; D013291:Streptococcus / Q000378:metabolism*; D027981:Symporters
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C050221:LacY protein, E coli; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; D027981:Symporters; C021499:lactose permease; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C030182:phosphocarrier protein HPr