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.
Norgaard-Nielsen
Kristine
K
Molecular Neuropharmacology Group, Department of Pharmacology 12-5-22, The Panum Institute, University of Copenhagen, DK-2200 N Copenhagen, Denmark.
Norregaard
Lene
L
Hastrup
Hanne
H
Javitch
Jonathan A
JA
Gether
Ulrik
U
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