Zn(2+) site engineering at the oligomeric interface of the dopamine transporter.
Increasing evidence suggests that Na(+)/Cl(-)-dependent neurotransmitter transporters exist as homo-oligomeric proteins. However, the functional implication of this oligomerization remains unclear. Here we demonstrate the engineering of a Zn(2+) binding site at the predicted dimeric interface of the dopamine transporter (DAT) corresponding to the external end of transmembrane segment 6. Upon binding to this site, which involves a histidine inserted in position 310 (V310H) and the endogenous Cys306 within the same DAT molecule, Zn(2+) potently inhibits [(3)H]dopamine uptake. These data provide indirect evidence that conformational changes critical for the translocation process may occur at the interface between two transporter molecules in the oligomeric structure.
Molecular Neuropharmacology Group, Department of Pharmacology 12-5-22, The Panum Institute, University of Copenhagen, DK-2200 N Copenhagen, Denmark.
Kristine
K
Norgaard-Nielsen
Lene
L
Norregaard
Hanne
H
Hastrup
Jonathan A
JA
Javitch
Ulrik
U
Gether
D000818:Animals; D001704:Biopolymers; D019556:COS Cells; D004298:Dopamine / Q000378:metabolism; D050483:Dopamine Plasma Membrane Transport Proteins; D006801:Humans; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D015032:Zinc / Q000737:chemistry / Q000378:metabolism*
D001704:Biopolymers; D050483:Dopamine Plasma Membrane Transport Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; C493214:SLC6A3 protein, human; D015032:Zinc; D004298:Dopamine