Publications

• Structure and mechanism of the glycerol-3-phosphate transporter from Escherichia coli.

  Huang Y., Lemieux M.J., Song J., Auer M., Wang D.-N.

  The major facilitator superfamily represents the largest group of secondary membrane transporters in the cell. Here we report the 3.3 angstrom resolution structure of a member of this superfamily, GlpT, which transports glycerol-3-phosphate into the cytoplasm and inorganic phosphate into the perip ... >> More

  The major facilitator superfamily represents the largest group of secondary membrane transporters in the cell. Here we report the 3.3 angstrom resolution structure of a member of this superfamily, GlpT, which transports glycerol-3-phosphate into the cytoplasm and inorganic phosphate into the periplasm. The amino- and carboxyl-terminal halves of the protein exhibit a pseudo two-fold symmetry. Closed off to the periplasm, a centrally located substrate-translocation pore contains two arginines at its closed end, which comprise the substrate-binding site. Upon substrate binding, the protein adopts a more compact conformation. We propose that GlpT operates by a single-binding site, alternating-access mechanism through a rocker-switch type of movement. << Less

  Science 301:616-620(2003) [PubMed] [EuropePMC]

• The pho regulon-dependent Ugp uptake system for glycerol-3-phosphate in Escherichia coli is trans inhibited by Pi.

  Brzoska P., Rimmele M., Brzostek K., Boos W.

  sn-Glycerol-3-phosphate (G3P) or glyceryl phosphoryl phosphodiesters, the substrates of the phoB-dependent Ugp transport system, when transported exclusively through this system, can serve as a sole source of phosphate but not as a sole source of carbon (H. Schweizer, M. Argast, and W. Boos, J. Ba ... >> More

  sn-Glycerol-3-phosphate (G3P) or glyceryl phosphoryl phosphodiesters, the substrates of the phoB-dependent Ugp transport system, when transported exclusively through this system, can serve as a sole source of phosphate but not as a sole source of carbon (H. Schweizer, M. Argast, and W. Boos, J. Bacteriol. 150:1154-1163, 1982). In order to explain this phenomenon, we tested two possibilities: repression of the pho regulon by Ugp-mediated transport and feedback inhibition by internal G3P or its degradation product Pi. Using an ugp-lacZ fusion, we found that the expression of ugp does not decline upon exposure to G3P, in contrast to the repressing effect of transport of Pi via the Pst system. This indicated that the Ugp system becomes inhibited after the uptake and metabolism of G3P. Using 32P-labeled G3P, we observed that little Pi is released by cells taking up G3P via the Ugp system but large amounts of Pi are released when the cells are taking up G3P via the GlpT system. Using a glpD mutant that could not oxidize G3P but which could still phosphorylate exogenous glycerol to G3P after GlpF-mediated transport of glycerol, we could not find trans inhibition of Ugp-mediated uptake of exogenous 14C-G3P. However, when allowing uptake of Pi via Pst, we observed a time-dependent inhibition of 14C-G3P taken up by the Ugp transport system. Inhibition was half maximal after 2 min and could be elicited by Pi concentrations below 0.5 mM. Cells had to be starved for Pi in order to observe this inhibition. We conclude that the activity of the Ugp transport system is controlled by the level of internal phosphate. << Less

  J. Bacteriol. 176:15-20(1994) [PubMed] [EuropePMC]

• Kinetic analysis by in vivo 31P nuclear magnetic resonance of internal Pi during the uptake of sn-glycerol-3-phosphate by the pho regulon-dependent Ugp system and the glp regulon-dependent GlpT system.

  Xavier K.B., Kossmann M., Santos H., Boos W.

  When sn-glycerol-3-phosphate (G3P) is taken up exclusively by the pho regulon-dependent Ugp transport system, it can be used as the sole source of Pi but not as the sole source of carbon. We had previously suggested that the inability of G3P to be used as a carbon source under these conditions is ... >> More

  When sn-glycerol-3-phosphate (G3P) is taken up exclusively by the pho regulon-dependent Ugp transport system, it can be used as the sole source of Pi but not as the sole source of carbon. We had previously suggested that the inability of G3P to be used as a carbon source under these conditions is due to trans inhibition of G3P uptake by internal Pi derived from the degradation of G3P (P. Brzoska, M. Rimmele, K. Brzostek, and W. Boos, J. Bacteriol. 176:15-20, 1994). Here we report 31P nuclear magnetic resonance measurements of intact cells after exposure to G3P as well as to Pi, using different mutants defective in pst (high-affinity Pi transport), ugp (pho-dependent G3P transport), glpT (glp-dependent G3P transport), and glpD (aerobic G3P dehydrogenase). When G3P was transported by the Ugp system and when metabolism of G3P was allowed (glpD+), Pi accumulated to about 13 to 19 mM. When G3P was taken up by the GlpT system, the preexisting internal Pi pool (whether low or high) did not change. Both systems were inversely controlled by internal Pi. Whereas the Ugp system was inhibited, the GlpT system was stimulated by elevated internal Pi. << Less

  J. Bacteriol. 177:699-704(1995) [PubMed] [EuropePMC]

• Effect of glpT and glpD mutations on expression of the phoA gene in Escherichia coli.

  Rao N.N., Roberts M.F., Torriani A., Yashphe J.

  In vivo 31P nuclear magnetic resonance analysis of Escherichia coli cells showed that the intracellular concentration of P(i) remained constant in wild-type and in a glpT mutant strain whether the cells were grown on excess (2 mM) P(i) or sn-glycerol-3-phosphate as a phosphate source. The function ... >> More

  In vivo 31P nuclear magnetic resonance analysis of Escherichia coli cells showed that the intracellular concentration of P(i) remained constant in wild-type and in a glpT mutant strain whether the cells were grown on excess (2 mM) P(i) or sn-glycerol-3-phosphate as a phosphate source. The function of the phoA promoter (measured by beta-galactosidase activity in a phoA-lacZ fusion strain) was repressed when glpT+ cells were utilizing sn-glycerol-3-phosphate as the sole source of phosphate. These cells were devoid of alkaline phosphatase activity. However, the phoA promoter was fully active in a glpT mutant. These results indicated that the repression of the enzyme synthesis was not due to a variation in the level of cytoplasmic P(i) but was due to the P(i) excreted into the periplasm and/or to the medium. << Less

  J Bacteriol 175:74-79(1993) [PubMed] [EuropePMC]