English
 
Help Privacy Policy Disclaimer
  Advanced SearchBrowse

Item

ITEM ACTIONSEXPORT

Released

Journal Article

High-affinity Cation Binding to Organic Cation Transporter OCT1 Induces Movement of Helix 11 and Blocks Transport after Mutations in a Modeled Interaction Domain between Two Helices

MPS-Authors
/persons/resource/persons137592

Bamberg,  Ernst
Department of Biophysical Chemistry, Max Planck Institute of Biophysics, Max Planck Society;

/persons/resource/persons137658

Friedrich,  Thomas
Department of Biophysical Chemistry, Max Planck Institute of Biophysics, Max Planck Society;

External Resource
No external resources are shared
Fulltext (restricted access)
There are currently no full texts shared for your IP range.
Fulltext (public)
There are no public fulltexts stored in PuRe
Supplementary Material (public)
There is no public supplementary material available
Citation

Gorbunov, D., Gorboulev, V., Shatskaya, N., Mueller, T., Bamberg, E., Friedrich, T., et al. (2008). High-affinity Cation Binding to Organic Cation Transporter OCT1 Induces Movement of Helix 11 and Blocks Transport after Mutations in a Modeled Interaction Domain between Two Helices. Molecular Pharmacology, 73(1), 50-61. doi: 10.1124/mol.107.040170.


Cite as: https://hdl.handle.net/11858/00-001M-0000-0024-D845-3
Abstract
Voltage-clamp fluorometry was performed with a cysteine-deprived mutant of rat organic cation transporter 1 (rOCT1) in which Phe483 in transmembrane alpha-helix (TMH) 11 close to the extracellular surface was replaced by cysteine and labeled with tetramethylrhodamine-6-maleimide. Potential-dependent fluorescence changes were observed that were sensitive to presence of substrates choline, tetraethylammonium (TEA), and 1-methyl-4-phenylpyridinium (MPP) and of the nontransported inhibitor tetrabutylammonium (TBuA). Using potential-dependent fluorescence changes as readout, one high-affinity binding site per substrate and two high-affinity binding sites for TBuA were identified in addition to the previously described single interaction sites. In a structure model of rOCT1 with an inward open cleft that was derived from a known crystal structure of lacY permease, Phe483 is close to Trp147 in TMH 2. In contrast, in a model with an outward open cleft these amino acids are far apart. After replacement of Phe483 or Trp147 by cysteine or serine, high-affinity binding of TBuA leads to inhibition of MPP or TEA uptake, whereas it has no effect on cation uptake by wild-type rOCT1. Coexisting high-affinity cation binding sites in organic cation transporters may collect low concentration xenobiotics and drugs; however, translocation including transitions between outward- and inward-oriented conformations may only be induced when a low-affinity cation binding site is loaded. We propose that cations bound to high-affinity sites may be translocated together with cations bound to low-affinity sites or that they may block the translocation mechanism.