Molecular Dynamics Approach to Determine the Importance of an Asparagine Residue on Chloride-Dependence in the Human Serotonin Transporter

The brain helps regulate all of our body's activities by maintaining a balance between active and inactive neurons. The majority of transporters involved in maintaining the brain's chemical balance can be grouped into a large gene family SLC6. Part of this family is responsible for transport of serotonin (SERT), dopamine (DAT), and norepenephrine (NET). The focus of our research is on the human serotonin transporter. The human serotonin transporter (hSERT) is of particular clinical significance as it is both a common target of psychostimulants, such as cocaine and MDMA (ecstasy), as well as a target for serotonin selective reuptake inhibitors (SSRI) used in the treatment of mood disorders.In our system, the crucial Na1 site is still present but some focus shifts to the Cl- site which is in near proximity. The exact function of the Cl- is unknown, but the human SERT cannot function without it. In our study we are utilizing mutagenesis, in particular of amino acids Asn101 and Ser336, to study why that may be. N101 is in close proximity to the ions as well as the substrate binding site, which makes it a good candidate for an ion coordinating residue. S336 functions as a partner to N101 in maintaining this chloride dependence. We utilized our wild-type, N101A, and S336C mutants to observe the coordination of the ions and 5HT. We observed the positioning of the ions/5HT in each of the mutants relative to the wild-type. We also calculated binding energies of each of the ions and the substrate. This will tell us the effect mutations have on the energetics of binding and coordination, as well as possible clue as to why selectivity and stoichiometry is affected.