Binding of Serotonin to the Human Serotonin Transporter. Molecular Modeling and Experimental Validation

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• 作者：Leyla Celik ; Steffen Sinning ; Kasper Severinsen ; Carsten G. Hansen ; Maria S. Mller ; Mikael Bols ; Ove Wiborg ; Birgit Schitt
• 刊名：Journal of the American Chemical Society
• 出版年：2008
• 出版时间：March 26, 2008
• 年：2008
• 卷：130
• 期：12
• 页码：3853 - 3865
• 全文大小：560K
• 年卷期：v.130,no.12(March 26, 2008)
• ISSN：1520-5126

文摘

Molecular modeling and structure-activity relationship studies were performed to propose amodel for binding of the neurotransmitter serotonin (5-HT) to the human serotonin transporter (hSERT).Homology models were constructed using the crystal structure of a bacterial homologue, the leucinetransporter from Aquifex aeolicus, as the template and three slightly different sequence alignments. Inducedfit docking of 5-HT into hSERT homology models resulted in two different binding modes. Both show a saltbridge between Asp98 and the charged primary amine of 5-HT, and both have the 5-HT C6 position of theindole ring pointing toward Ala173. The difference between the two orientations of 5-HT is an enantiofacialdiscrimination of the indole ring, resulting in the 5-hydroxyl group of 5-HT being vicinal to either Ser438/Thr439 or Ala169/Ile172/Ala173. To assess the binding experimentally, binding affinities for 5-HT and 17analogues toward wild type and 13 single point mutants of hSERT were measured using an approachtermed paired mutant-ligand analogue complementation (PaMLAC). The proposed ligand-proteininteraction was systematically examined by disrupting it through site-directed mutagenesis and re-establishing another interaction via a ligand analogue matching the mutated residue, thereby minimizingthe risk of identifying indirect effects. The interactions between Asp98 and the primary amine of 5-HT andthe interaction between the C6-position of 5-HT and hSERT position 173 was confirmed using PaMLAC.The measured binding affinities of various mutants and 5-HT analogues allowed for a distinction betweenthe two proposed binding modes of 5-HT and biochemically support the model for 5-HT binding in hSERTwhere the 5-hydroxyl group is in close proximity to Thr439.