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Nonetheless, at the S2 site, methylphenidate exhibited an interplay pattern and binding orientation more akin to that of modafinil-forming a cation-π interplay between the ligand aromatic ring and R85, with the protonated ligand amine anchored by a combination of hydrogen bonding with D476 and a cation-π interaction with the aromatic facet chain of F320 (Figs. As well as, the 3β-fluorophenyl ring of CFT participated in π-π stacking aromatic interaction with the side-chain of F326 and the 2β-carbomethoxy moiety formed a hydrogen bond with S422 of TM8 (Figs. The space between the carboxylate oxygen atom of D79 and the ring hydroxyl moiety of Y156 is displayed within the lower right of every panel (in yellow). Binding of the atypical inhibitor benztropine, nevertheless, resulted in a preserved D79-Y156 hydrogen bond (i.e. an interatomic distance less than 3.5 Å), suggesting that-in contrast to cocaine-binding of benztropine at the S1 site doesn't prevent closure of the gate. 2008), docking of β-CFT at the S1 site resulted in a D79-Y156 distance of 4.85 Å, indicative of an open extracellular gate (Fig. 5A). Apparently, at the S2 site, extension of CFT's 3β-fluorophenyl moiety downward into the S1 site permitted an aromatic stacking interplay with Y156, pushing the tyrosine ring aside and increasing the D79-Y156 distance to 4.94 Å (Fig. 5B). As well as, the classical inhibitor d-methylphenidate also disrupted the D79-Y156 hydrogen bond, yielding S1- and S2-bound distances of 4.12 Å and 3.57 Å, respectively (Fig. 5C-D). This means that cocaine-like phenyltropane inhibitors and methylphenidate are capable of inducing an open-to-out transporter conformation upon binding at both the S2 or S1 site.

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