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- Title
Impact of disruption of secondary binding site S<sub>2</sub> on dopamine transporter function.
- Authors
Zhen, Juan; Reith, Maarten E. A.
- Abstract
The structures of the leucine transporter, drosophila dopamine transporter, and human serotonin transporter show a secondary binding site (designated S2) for drugs and substrate in the extracellular vestibule toward the membrane exterior in relation to the primary substrate recognition site (S1). The present experiments are aimed at disrupting S2 by mutating Asp476 and Ile159 to Ala. Both mutants displayed a profound decrease in [3H] DA uptake compared with wild-type associated with a reduced turnover rate kcat. This was not caused by a conformational bias as the mutants responded to Zn2+ (10 μM) similarly as WT. The dopamine transporters with either the D476A or I159A mutation both displayed a higher Ki for dopamine for the inhibition of [3H](−)-2-β-carbomethoxy-3-β-(4-fluorophenyl)tropane binding than did the WT transporter, in accordance with an allosteric interaction between the S1 and S2 sites. The results provide evidence in favor of a general applicability of the two-site allosteric model of the Javitch/Weinstein group from LeuT to dopamine transporter and possibly other monoamine transporters.
- Subjects
LEUCINE; SEROTONIN transporters; DOPAMINE; TROPANES; GLUCOPYRANOSIDE
- Publication
Journal of Neurochemistry, 2016, Vol 138, Issue 5, p694
- ISSN
0022-3042
- Publication type
Article
- DOI
10.1111/jnc.13704