Depression, anxiety disorders, ADHD, and insomnia can result from malfunctions of the serotonin transporter (SERT). Computational models of drug binding show that there are two possible ways a drug may interact with SERT. In addition, lipid interactions with SERT may affect the drug’s binding affinity and efficacy. By investigating drug binding and lipid interactions via photocrosslinking, a more accurate model can be developed for better drug design. Human SERT (hSERT) was purified from TREX after tetracycline induction. A benzophenone derivatized fluoxetine was photocrosslinked with purified hSERT. After trypsinolysis, mass shifted crosslinks were identified using quadrupole time-of-flight mass spectrometry (QTOF) to resolve its binding pocket. Similarly, pacFA-18:1 phosphatidylcholine was incorporated into reconstituted hSERT vesicles and subjected to similar MS analysis. The resulting state-dependent crosslinks may be used to map lipid-accessible regions of hSERT in different allosteric states.

Structural analysis of the serotonin transporter via crosslinking and peptide mass fingerprinting

251st National Meeting (2016)

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