Journal of Neuroscience, Vol 9, 2664-2670, Copyright © 1989 by Society for Neuroscience
Dopamine transport sites selectively labeled by a novel photoaffinity probe: 125I-DEEP
DE Grigoriadis, AA Wilson, R Lew, JS Sharkey and MJ Kuhar
Neuroscience Branch, National Institute on Drug Abuse, Baltimore, Maryland 21224.
The dopamine transporter was labeled using a photosensitive compound
related to GBR-12909, 125I-1-[2-(diphenylmethoxy)ethyl]-4-[2- (4-azido-
3-iodophenyl)ethyl]piperazine (125I-DEEP). 125I-DEEP bound reversibly and
with high affinity to the dopamine transport protein in the absence of
light and could be covalently attached to the protein following exposure to
UV light. In rat striatal homogenates, 125I-DEEP was found to incorporate
covalently into a protein with apparent molecular weight of 58,000 Da. The
properties of this binding protein were characteristic of the dopamine
transporter since covalent attachment could be inhibited by dopamine-uptake
blockers with the proper pharmacological rank order of potencies. Covalent
binding was also inhibited in a stereospecific manner by (+) and (-)
cocaine, as well as other cocaine analogs. The protein was not found in the
cerebellum. The dopamine transporter appears to exist in a glycosylated
form since photoaffinity-labeled transport sites could adsorb to wheat
germ- agglutinin and could be specifically eluted from the column by
beta-N- acetylglucosamine.
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