title: Fluorescent stilbazolium dyes as probes of the norepinephrine transporter: structural insights into substrate binding.
abstract: We report the synthesis, binding kinetics, optical spectroscopy and predicted binding modes of a series of sterically demanding, fluorescent norepinephrine transporter (NET) ligands. A series of bulky stilbazolium dyes, including six newly synthesized compounds, were evaluated to determine the effect of extending the molecular probes'    'heads' or 'tails'. Taking advantage of the dyes' characteristic 'turn-on' emission, the kinetic binding parameters, k(on) and k(off) were determined revealing that extension of the molecules' tails is well tolerated while expansion of the head is not. Additionally, a 'headfirst' orientation appears to be preferred over a 'tail-first' binding pose. Further details of the possible binding modes were obtained from the emission spectra of the bound probes. A small range of interplanar twist angles, approximately 35° to 60°, is predicted to produce the observed emission. Docking experiments and molecular modelling support the kinetic and spectroscopic data providing structural insights into substrate binding.
authors.affiliation: Department of Chemistry, University of Miami, 1301 Memorial Drive, Coral Gables, FL 33124, USA. jnwilson@miami.edu
authors.forename: James N
authors.initials: JN
authors.lastname: Wilson
authors.forename: Adrienne S
authors.initials: AS
authors.lastname: Brown
authors.forename: W Michael
authors.initials: WM
authors.lastname: Babinchak
authors.forename: Clark D
authors.initials: CD
authors.lastname: Ridge
authors.forename: Jamie D
authors.initials: JD
authors.lastname: Walls
mesh_terms: D001665:Binding Sites; D002478:Cells, Cultured; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D008024:Ligands; D018613:Microscopy, Confocal; D008958:Models, Molecular; D015335:Molecular Probes / Q000138:chemical synthesis / Q000737:chemistry*; D015394:Molecular Structure; D050484:Norepinephrine Plasma Membrane Transport Proteins / Q000737:chemistry*; D011726:Pyridinium Compounds / Q000138:chemical synthesis / Q000737:chemistry*; D011789:Quantum Theory
chemical_list: C032315:4-(4-dimethylaminostyryl)-1-methylpyridinium; D005456:Fluorescent Dyes; D008024:Ligands; D015335:Molecular Probes; D050484:Norepinephrine Plasma Membrane Transport Proteins; D011726:Pyridinium Compounds