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Our main goal was to investigate the pathogenic mechanisms of this variant, located near the neurotransmitter permeation pathway, and compare it with other variants located either in the permeation pathway or close to the lipid bilayer. The mutants G307R and A334P, close to the gates of the transporter, could be glycosylated with variable efficiency and reached the membrane, albeit inactive. Mutants located in the center of the permeation pathway (G297R) or close to the lipid bilayer (A128V, G550R) were retained in the endoplasmic reticulum. Applying an Elastic Network Model, to these and to other previously characterized variants, we found that G307R and A334P significantly perturb the structure and dynamics of the intracellular gate, which can explain their reduced activity, while for A228V and G362R, the reduced translocation to the membrane quantitatively accounts for the reduced activity. The addition of a chemical chaperone (4-phenylbutyric acid, PBA), which improves protein folding, increased the activity of GAT-1WT, as well as most of the assayed variants, including G307R, suggesting that PBA might also assist the conformational changes occurring during the alternative access transport cycle.<\/jats:p>","DOI":"10.3390\/ijms24020955","type":"journal-article","created":{"date-parts":[[2023,1,5]],"date-time":"2023-01-05T03:12:15Z","timestamp":1672888335000},"page":"955","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Experimental and Bioinformatic Insights into the Effects of Epileptogenic Variants on the Function and Trafficking of the GABA Transporter GAT-1"],"prefix":"10.3390","volume":"24","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1158-0226","authenticated-orcid":false,"given":"Dolores","family":"Piniella","sequence":"first","affiliation":[{"name":"Centro de Biolog\u00eda Molecular Severo Ochoa, Facultad de Ciencias, Consejo Superior de Investigaciones Cient\u00edficas, Universidad Aut\u00f3noma de Madrid, 28049 Madrid, Spain"},{"name":"IdiPAZ, Institute of Health Carlos III (ISCIII), 28046 Madrid, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7622-5722","authenticated-orcid":false,"given":"Ania","family":"Canseco","sequence":"additional","affiliation":[{"name":"Centro de Biolog\u00eda Molecular Severo Ochoa, Facultad de Ciencias, Consejo Superior de Investigaciones Cient\u00edficas, Universidad Aut\u00f3noma de Madrid, 28049 Madrid, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Silvia","family":"Vidal","sequence":"additional","affiliation":[{"name":"Sant Joan de D\u00e9u Research Foundation, 08950 Barcelona, Spain"},{"name":"IPER, Institut de Recerca Pedi\u00e0trica Hospital Sant Joan de D\u00e9u, 08950 Barcelona, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Clara","family":"Xiol","sequence":"additional","affiliation":[{"name":"Sant Joan de D\u00e9u Research Foundation, 08950 Barcelona, Spain"},{"name":"IPER, Institut de Recerca Pedi\u00e0trica Hospital Sant Joan de D\u00e9u, 08950 Barcelona, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ar\u00e1nzazu","family":"D\u00edaz de Bustamante","sequence":"additional","affiliation":[{"name":"Genetics Unit, Hospital Universitario de M\u00f3stoles, 28935 Madrid, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0578-9568","authenticated-orcid":false,"given":"Itxaso","family":"Mart\u00ed-Carrera","sequence":"additional","affiliation":[{"name":"Pediatric Neurology, Hospital Universitario Donostia, Biodonostia, Universidad del Pa\u00eds Vasco UPV-EHU, 20014 San Sebasti\u00e1n, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0588-9307","authenticated-orcid":false,"given":"Judith","family":"Armstrong","sequence":"additional","affiliation":[{"name":"IPER, Institut de Recerca Pedi\u00e0trica Hospital Sant Joan de D\u00e9u, 08950 Barcelona, Spain"},{"name":"Molecular and Genetics Medicine Section, Hospital Sant Joan de D\u00e9u, 08950 Barcelona, Spain"},{"name":"CIBER-ER (Biomedical Network Research Center for Rare Diseases), Institute of Health Carlos III (ISCIII), 28029 Madrid, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ugo","family":"Bastolla","sequence":"additional","affiliation":[{"name":"Centro de Biolog\u00eda Molecular Severo Ochoa, Facultad de Ciencias, Consejo Superior de Investigaciones Cient\u00edficas, Universidad Aut\u00f3noma de Madrid, 28049 Madrid, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Francisco","family":"Zafra","sequence":"additional","affiliation":[{"name":"Centro de Biolog\u00eda Molecular Severo Ochoa, Facultad de Ciencias, Consejo Superior de Investigaciones Cient\u00edficas, Universidad Aut\u00f3noma de Madrid, 28049 Madrid, Spain"},{"name":"IdiPAZ, Institute of Health Carlos III (ISCIII), 28046 Madrid, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2023,1,4]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"808","DOI":"10.1016\/j.ajhg.2015.02.016","article-title":"Mutations in the GABA Transporter SLC6A1 Cause Epilepsy with Myoclonic-Atonic Seizures","volume":"96","author":"Carvill","year":"2015","journal-title":"Am. 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