Zn(2+) site engineering at the oligomeric interface of the dopamine transporter

Kristine Norgaard-Nielsen; Lene Norregaard; Hanne Hastrup; Jonathan A Javitch; Ulrik Gether

Zn(2+) site engineering at the oligomeric interface of the dopamine transporter

Kristine Norgaard-Nielsen, Lene Norregaard, Hanne Hastrup, Jonathan A Javitch, Ulrik Gether

33 Citations (Scopus)

Abstract

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.

Original language	English
Journal	FEBS Letters
Volume	524
Issue number	1-3
Pages (from-to)	87-91
Number of pages	5
ISSN	0014-5793
Publication status	Published - 31 Jul 2002

Keywords

Animals
Biopolymers
COS Cells
Dopamine
Dopamine Plasma Membrane Transport Proteins
Humans
Membrane Glycoproteins
Membrane Transport Proteins
Mutagenesis, Site-Directed
Nerve Tissue Proteins
Polymerase Chain Reaction
Protein Binding
Zinc

Cite this

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title = "Zn(2+) site engineering at the oligomeric interface of the dopamine transporter",

abstract = "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.",

keywords = "Animals, Biopolymers, COS Cells, Dopamine, Dopamine Plasma Membrane Transport Proteins, Humans, Membrane Glycoproteins, Membrane Transport Proteins, Mutagenesis, Site-Directed, Nerve Tissue Proteins, Polymerase Chain Reaction, Protein Binding, Zinc",

author = "Kristine Norgaard-Nielsen and Lene Norregaard and Hanne Hastrup and Javitch, {Jonathan A} and Ulrik Gether",

year = "2002",

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language = "English",

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TY - JOUR

T1 - Zn(2+) site engineering at the oligomeric interface of the dopamine transporter

AU - Norgaard-Nielsen, Kristine

AU - Norregaard, Lene

AU - Hastrup, Hanne

AU - Javitch, Jonathan A

AU - Gether, Ulrik

PY - 2002/7/31

Y1 - 2002/7/31

N2 - 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.

AB - 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.

KW - Animals

KW - Biopolymers

KW - COS Cells

KW - Dopamine

KW - Dopamine Plasma Membrane Transport Proteins

KW - Humans

KW - Membrane Glycoproteins

KW - Membrane Transport Proteins

KW - Mutagenesis, Site-Directed

KW - Nerve Tissue Proteins

KW - Polymerase Chain Reaction

KW - Protein Binding

KW - Zinc

M3 - Journal article

C2 - 12135746

SN - 0014-5793

VL - 524

SP - 87

EP - 91

JO - F E B S Letters

JF - F E B S Letters

IS - 1-3

ER -

Zn(2+) site engineering at the oligomeric interface of the dopamine transporter

Abstract

Keywords

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