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oai:figshare.com:article/32641791 2026-10-01T16:20:34Z portal_1172 item_type_8 month_year_10_2026

Structure-function studies of Borrelia metal transport A (BmtA) from Lyme disease-causing Borrelia burgdorferi Raja Massoud Ali Alawar (24170250) PUREID: 656864683 BmtA Lyme disease Transmembrane metal transport Lyme disease, caused by the spirochetal bacterium Borrelia burgdorferi, remains the most prevalent vector-borne illness in humans. Addressing challenges such as antibiotic resistance, persistent infection, and complex disease dynamics requires identifying novel therapeutic targets. Manganese (Mn²˖) is essential for the survival of burgdorferi, serving as a cofactor for enzymes that protect against oxidative stress, particularly superoxide dismutase. The Borrelia metal transporter A (BmtA), a membra protein responsible for Mn²˖ uptake, is critical for bacterial infectivity, making it a promising drug target. This study characterized the structural and functional properties BmtA to elucidate its role in manganese transport. A structural model was generated using AlphaFold2 and compared with bacterial and mammalian ZIP transporters to identify metal-binding residues. Site-directed mutagenesis and functional analyses in a yeast expression system, combined with PAR metal-binding and ICP-MS assays, identified residues H133, E137, and E203 as essential for Mn²˖ transport. Zinc was also found to stabilize BmtA, with deficiency resulting in protein degradation.<br><br>For biochemical characterization, optimal solubilization and purification were achieved using n-Dodecyl-β-D-Maltopyranoside (DDM), maintaining the protein’s dimeric stat as confirmed by Blue Native PAGE and crosslinking experiments. Metal-binding and thermal shift assays demonstrated that Mn²˖ induced conformational stabilization, whereas Zn²˖ had minimal impact. Mutations at key residues disrupted folding and stability, highlighting the importance of metal coordination.Overall, this study enhance understanding of BmtA’s structure–function relationship and supports its potential as a therapeutic target for combating Lyme disease.<br><br><i>Thesis is embargoed until 31 December 2026</i>. 2026-10-01T16:20:34Z Text Thesis 10.17034/32641791.v1 https://figshare.com/articles/thesis/Structure-function_studies_of_Borrelia_metal_transport_A_BmtA_from_Lyme_disease-causing_Borrelia_burgdorferi/32641791 All Rights Reserved Open Access after 2026-12-31