Asp22 drives the protonation state of the Staphylococcus epidermidis glucose/H+ symporter.
The Staphylococcus epidermidis glucose/H+ symporter (GlcPSe) is a membrane transporter highly specific for glucose and a homolog of the human glucose transporters (GLUT, SLC2 family). Most GLUTs and their bacterial counterparts differ in the transport mechanism, adopting uniport and sugar/H+ symport, respectively. Unlike other bacterial GLUT homologs (for example, XylE), GlcPSe has a loose H+/sugar coupling. Asp22 is part of the proton-binding site of GlcPSe and crucial for the glucose/H+ co-transport mechanism. To determine how pH variations affect the proton site and the transporter, we performed surface-enhanced IR absorption spectroscopy on the immobilized GlcPSe We found that Asp22 has a pK of 8.5 ± 0.1, a value consistent with that determined previously for glucose transport, confirming the central role of this residue for the transport mechanism of GlcPSe A neutral replacement of the negatively charged Asp22 led to positive charge displacements over the entire pH range, suggesting that the polarity change of the WT reflects the protonation state of Asp22 We expected that the substitution of the residue Ile105 for a serine, located within hydrogen-bonding distance to Asp22, would change the microenvironment, but the pK of Asp22 corresponded to that of the WT. A167E mutation, selected in analogy to the XylE, introduced an additional protonatable site and perturbed the protonation state of Asp22, with the latter now exhibiting a pK of 6.4. These studies confirm that Asp22 is the proton-binding residue in GlcPSe and show that charged residues in its vicinity affect the pK of glucose/H+ symport.
['Seica|Ana Filipa Santos|AFS|', 'Iancu|Cristina V|CV|', 'Pfeilschifter|Benedikt|B|', 'Madej|M Gregor|MG|', 'Choe|Jun-Yong|JY|', 'Hellwig|Petra|P|']
D001224:Aspartic Acid D001692:Biological Transport D005947:Glucose D051246:Glucose Transport Proteins, Facilitative D006863:Hydrogen-Ion Concentration D011522:Protons D013212:Staphylococcus epidermidis D027981:Symporters
membrane transport transport glucose transport spectroscopy major facilitator family major facilitator superfamily membrane proteins pK value proton transport surface-enhanced infrared spectroscopy transport proteins