Molecular determinants of Neu5Ac binding to a tripartite ATP independent periplasmic (TRAP) transporter
N -Acetylneuraminic acid (Neu5Ac) is a negatively charged nine-carbon amino sugar that is often the peripheral sugar in human cell-surface glycoconjugates. Some bacteria scavenge, import, and metabolize Neu5Ac or redeploy it on their cell surfaces for immune evasion. The import of Neu5Ac by many bac...
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eLife Sciences Publications Ltd
2025-02-01
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| Online Access: | https://elifesciences.org/articles/98158 |
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| author | Parveen Goyal KanagaVijayan Dhanabalan Mariafrancesca Scalise Rosmarie Friemann Cesare Indiveri Renwick CJ Dobson Kutti R Vinothkumar Subramanian Ramaswamy |
| author_facet | Parveen Goyal KanagaVijayan Dhanabalan Mariafrancesca Scalise Rosmarie Friemann Cesare Indiveri Renwick CJ Dobson Kutti R Vinothkumar Subramanian Ramaswamy |
| author_sort | Parveen Goyal |
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| description | N -Acetylneuraminic acid (Neu5Ac) is a negatively charged nine-carbon amino sugar that is often the peripheral sugar in human cell-surface glycoconjugates. Some bacteria scavenge, import, and metabolize Neu5Ac or redeploy it on their cell surfaces for immune evasion. The import of Neu5Ac by many bacteria is mediated by tripartite ATP-independent periplasmic (TRAP) transporters. We have previously reported the structures of SiaQM, a membrane-embedded component of the Haemophilus influenzae TRAP transport system, (Currie et al., 2024). However, none of the published structures contain Neu5Ac bound to SiaQM. This information is critical for defining the transport mechanism and for further structure-activity relationship studies. Here, we report the structures of Fusobacterium nucleatum SiaQM with and without Neu5Ac. Both structures are in an inward (cytoplasmic side) facing conformation. The Neu5Ac-bound structure reveals the interactions of Neu5Ac with the transporter and its relationship with the Na+ binding sites. Two of the Na+-binding sites are similar to those described previously. We identify a third metal-binding site that is further away and buried in the elevator domain. Ser300 and Ser345 interact with the C1-carboxylate group of Neu5Ac. Proteoliposome-based transport assays showed that Ser300-Neu5Ac interaction is critical for transport, whereas Ser345 is dispensable. Neu5Ac primarily interacts with residues in the elevator domain of the protein, thereby supporting the elevator with an operator mechanism. The residues interacting with Neu5Ac are conserved, providing fundamental information required to design inhibitors against this class of proteins. |
| format | Article |
| id | doaj-art-2cdbb9853d2e49f6b949626d240830e1 |
| institution | Kabale University |
| issn | 2050-084X |
| language | English |
| publishDate | 2025-02-01 |
| publisher | eLife Sciences Publications Ltd |
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| series | eLife |
| spelling | doaj-art-2cdbb9853d2e49f6b949626d240830e12025-02-06T15:41:19ZengeLife Sciences Publications LtdeLife2050-084X2025-02-011310.7554/eLife.98158Molecular determinants of Neu5Ac binding to a tripartite ATP independent periplasmic (TRAP) transporterParveen Goyal0https://orcid.org/0000-0002-7808-8298KanagaVijayan Dhanabalan1https://orcid.org/0000-0001-8636-2616Mariafrancesca Scalise2Rosmarie Friemann3Cesare Indiveri4Renwick CJ Dobson5https://orcid.org/0000-0002-5506-4939Kutti R Vinothkumar6https://orcid.org/0000-0002-6746-5684Subramanian Ramaswamy7https://orcid.org/0000-0002-6709-190XBiochemical Sciences Division, CSIR-National Chemical Laboratory, Pune, India; Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, India; Institute for Stem Cell Science and Regenerative Medicine, Bengaluru, IndiaBiological Sciences, Purdue University, West Lafayette, United StatesDepartment DiBEST (Biologia, Ecologia, Scienze della Terra) Unit of Biochemistry and Molecular Biotechnology, University of Calabria, Arcavacata di Rende, ItalyCentre for Antibiotic Resistance Research (CARe) at University of Gothenburg, Gothenburg, SwedenDepartment DiBEST (Biologia, Ecologia, Scienze della Terra) Unit of Biochemistry and Molecular Biotechnology, University of Calabria, Arcavacata di Rende, Italy; CNR, Institute of Biomembranes, Bioenergetics and Molecular Biotechnologies (IBIOM), via Amendola, Bari, ItalyBiomolecular Interaction Centre, Maurice Wilkins Centre for Biodiscovery, MacDiarmid Institute for Advanced Materials and Nanotechnology, and School of Biological Sciences, University of Canterbury, Christchurch, New Zealand; Department of Biochemistry and Pharmacology, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, Melbourne, AustraliaNational Centre for Biological Sciences TIFR, GKVK Campus, Bellary Road, Bengaluru, IndiaBiological Sciences, Purdue University, West Lafayette, United StatesN -Acetylneuraminic acid (Neu5Ac) is a negatively charged nine-carbon amino sugar that is often the peripheral sugar in human cell-surface glycoconjugates. Some bacteria scavenge, import, and metabolize Neu5Ac or redeploy it on their cell surfaces for immune evasion. The import of Neu5Ac by many bacteria is mediated by tripartite ATP-independent periplasmic (TRAP) transporters. We have previously reported the structures of SiaQM, a membrane-embedded component of the Haemophilus influenzae TRAP transport system, (Currie et al., 2024). However, none of the published structures contain Neu5Ac bound to SiaQM. This information is critical for defining the transport mechanism and for further structure-activity relationship studies. Here, we report the structures of Fusobacterium nucleatum SiaQM with and without Neu5Ac. Both structures are in an inward (cytoplasmic side) facing conformation. The Neu5Ac-bound structure reveals the interactions of Neu5Ac with the transporter and its relationship with the Na+ binding sites. Two of the Na+-binding sites are similar to those described previously. We identify a third metal-binding site that is further away and buried in the elevator domain. Ser300 and Ser345 interact with the C1-carboxylate group of Neu5Ac. Proteoliposome-based transport assays showed that Ser300-Neu5Ac interaction is critical for transport, whereas Ser345 is dispensable. Neu5Ac primarily interacts with residues in the elevator domain of the protein, thereby supporting the elevator with an operator mechanism. The residues interacting with Neu5Ac are conserved, providing fundamental information required to design inhibitors against this class of proteins.https://elifesciences.org/articles/98158TRAP transporterNeu5Ac bindingSodium ionsialic acid transportFusobacteriumcryo-EM |
| spellingShingle | Parveen Goyal KanagaVijayan Dhanabalan Mariafrancesca Scalise Rosmarie Friemann Cesare Indiveri Renwick CJ Dobson Kutti R Vinothkumar Subramanian Ramaswamy Molecular determinants of Neu5Ac binding to a tripartite ATP independent periplasmic (TRAP) transporter eLife TRAP transporter Neu5Ac binding Sodium ion sialic acid transport Fusobacterium cryo-EM |
| title | Molecular determinants of Neu5Ac binding to a tripartite ATP independent periplasmic (TRAP) transporter |
| title_full | Molecular determinants of Neu5Ac binding to a tripartite ATP independent periplasmic (TRAP) transporter |
| title_fullStr | Molecular determinants of Neu5Ac binding to a tripartite ATP independent periplasmic (TRAP) transporter |
| title_full_unstemmed | Molecular determinants of Neu5Ac binding to a tripartite ATP independent periplasmic (TRAP) transporter |
| title_short | Molecular determinants of Neu5Ac binding to a tripartite ATP independent periplasmic (TRAP) transporter |
| title_sort | molecular determinants of neu5ac binding to a tripartite atp independent periplasmic trap transporter |
| topic | TRAP transporter Neu5Ac binding Sodium ion sialic acid transport Fusobacterium cryo-EM |
| url | https://elifesciences.org/articles/98158 |
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