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Title: Structure modelling of gamma-aminobutyric acid and glycine transporters as important biological targets
Abstract:
Gamma-aminobutyric acid (GABA) and glycine are two primary inhibitory neurotransmitters in the central nervous system (CNS). In addition, glycine influences the activity of the glutamatergic system by acting as a co-agonist of the NMDA receptor. One of the main regulators of GABA and glycine concentration in the CNS are the membrane transporters belonging to the SLC6 family. There are 4 types of GABA transporters: GAT-1, BGT-1, GAT-2, GAT-3; and two main types of glycine transporters: GlyT-1, GlyT-2. Studies indicate that blocking GABA and glycine reuptake may be beneficial in the treatment of many disorders, including epilepsy, anxiety, depression, neuropathic pain, schizophrenia, and Alzheimer's disease. Although the potential for the use of GABA and glycine transporter inhibitors appears to be high, the only drug approved for treatment to date is tiagabine, a selective GAT-1 inhibitor used in the therapy of refractory epilepsy. One of the reasons for the failure to design new inhibitors with the desired activity, selectivity, and physicochemical properties is the lack of knowledge about the exact structure of particular types of GABA and glycine transporters, especially about their binding sites and mode of interaction with ligands. Therefore, the overall objective of this doctoral dissertation was to determine the exact structure of the transporters for gamma-aminobutyric a ; cid (GAT-1, BGT-1, GAT-2, GAT-3) and glycine (GlyT-1, GlyT-2) using molecular modelling methods. In the course of the study, a diverse pool of homology models of each transporter was generated. From this pool, the final models were selected which, according to the assessment of the applied bioinformatic tools, best reflected the actual structure of the proteins and explained the structure-activity relationship for the ligands docked into their binding sites. The constructed models allowed for a comparative analysis of the spatial structure of all types of GABA and glycine transporters, along with a comparison with other related proteins of known structure. The analysis focused on the structure of the binding sites, including the identification of the amino acids that have the greatest influence on the properties of specific transporters. The indicated differences were reflected in the results of the molecular docking of the reference ligands of each transporter. The stability of the binding modes obtained in the docking studies was verified by molecular dynamics simulations. Additionally, in some analyses, calculations of the free energy of ligand binding were carried out to indicate a more favourable binding mode. In silico studies show that the carboxyl group of GABA transporter inhibitors with amino acid structure is located at the main binding site (S1), while the aromatic ; rings of the compounds reach the S2 site within the vestibule of the transporter in the outward open state. The binding mode of the inhibitors explains the differences in the activity of their isomers, as well as the selectivity toward particular transporter types. Inhibitors with a non-amino acid structure, including novel 4-aminobutanamide and 4-hydroxybutanamide derivatives as well as their analogues obtained in the Department of Physicochemical Drug Analysis JU MC, bind mainly within the vestibule of GABA transporters. Fragments that may account for the slightly different arrangement of selected derivatives in particular types of GABA transporters and contribute to their varied activity were identified. In the case of GlyT-1, competitive inhibitors are arranged coherently at the S1 and S2 sites of the transporters. In contrast, non-competitive inhibitors are located partially on the intracellular side of the transporter in the inward open state. This binding mode is consistent with that observed in the recently released GlyT-1 crystal structure. In the case of GlyT-2, the currently known ligands bind at the S1 and/or S2 site of the transporter in the outward open state. The amino acid differences between GlyT-1 and GlyT-2 provide a reasonable explanation for the selectivity of the particular inhibitors. A virtual screening of the ZINC15 database for new BGT-1 transporter in ; hibitors was also carried out as a part of this thesis. The screening was based on the physicochemical properties of the compounds, a simplified pharmacophore model and molecular docking to the BGT-1 homology models. Finally, 12 compounds were selected, purchased and subsequently tested in vitro in the [3H]GABA uptake assay. The most active compound KL-202 shows moderate potency on the BGT-1 (IC50 = 48.9 μM) and GAT-3 (IC50 = 46.8 μM) transporters. The low molecular weight and high synthetic availability of compound KL-202 provide the potential for further development to obtain derivatives with higher activity and selectivity. The research carried out in this doctoral dissertation significantly broadened the knowledge of the structure of GABA and glycine transporters. This can support the rational design of their new inhibitors with desirable properties.
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Rada Dyscypliny Nauki farmaceutyczne
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Last modified:
Dec 9, 2024
In our library since:
Apr 8, 2024
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4
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http://dl.cm-uj.krakow.pl:8080/publication/5063
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| Edition name | Date |
|---|---|
| ZB-139427 | Dec 9, 2024 |
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