The top 20 dopamine transporter mutants: structure-function relationships and cocaine actions.

Our laboratory and others elucidated the primary amino acid sequences of the dopamine transporter (DAT) by cloning its cDNA and genomic sequences more than 12 years ago. Motivations for this work included the ideas that cocaine's interactions with DAT accounted for its rewarding properties and that selective inhibitors of DAT/cocaine interactions might thus provide good anticocaine medications. Such ideas supported interest in the detailed structure-function relationships of cocaine/DAT interactions, and in the construction and characterization of extensive series of site-directed DAT mutants. We can now select the most interesting 20 cocaine-analog selective mutations of the more than 100 single- and multiple amino acid substitution mutations that we have characterized. These mutants selectively reduce the affinities of the mutant DATs for cocaine analogs, but (absolutely or relatively) spare their affinities for dopamine. Several themes relevant to cocaine/DAT interactions emerge from these mutants. First, such mutations are found in a number of different DAT domains. Secondly, many but not all of these mutations lie in groups, near each other and near the same faces of presumably helical DAT transmembrane domains. Third, most are also conserved in the serotonin transporter (SERT), a transporter that is now strongly implicated in cocaine reward based on data from knockout mice. We discuss the results from these "top 20" mutants in light of the strengths and limitations of current DAT models and data from other studies. Taken together, these studies appear to indicate direct or indirect participation of several specific portions of DAT in selective recognition of cocaine analogs. These studies provide a strong basis for redirected studies aimed at producing dopamine- and serotonin-sparing cocaine antagonists that would represent combined DAT/SERT disinhibitors.