Poster Presentation The 48th Lorne Conference on Protein Structure and Function 2023

When you are an à la carte eater in a buffet world: Understanding selective bacterial nutrient uptake through structural and functional analysis (#336)

Michael C Newton-Vesty 1 2 , Joshua D Wright 1 2 , Michael J Currie 1 2 , James S Davies 3 , Gayan S Abeysekera 1 2 , Peter D Mace 4 5 , Rachel A North 3 , Renwick CJ Dobson 1 2 6
  1. S, University of Canterbury, Christchurch, Canterbury, New Zealand
  2. Biomolecular Interaction Centre and School of Biological Sciences, University of Canterbury, Christchurch, Canterbury, New Zealand
  3. Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
  4. Biochemistry Department and School of Biomedical Sciences, University of Otago, Dunedin, Otago, New Zealand
  5. S, University of Otago, Dunedin, Otago, New Zealand
  6. Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Melbourne, Victoria, Australia

Although the environment presents a smorgasbord of essential-to-life nutrients, it is the ability to select specific nutrients for import that enables most living organisms to thrive. One family of membrane imbedded proteins that accomplish nutrient import are ATP-independent periplasmic (TRAP) transporters, commonly found throughout bacteria and archaea. This family is known for selective transport of carboxylate and sulfonate containing nutrients and are the only known secondary transporters (not requiring ATP) that are dependent on an auxiliary substrate binding protein. This substrate binding protein “P” is located free in the periplasm (or tethered to the outer membrane) and sequesters the substrate before delivering it to the membrane bound components, “Q” and “M”, for transport into the cell. Recent studies have elucidated the structure of the membrane component of the transporter family, shedding light on the import mechanism. However, questions remain around how the substrate binding protein interacts with the membrane components to form the full transport cycle.

I am investigating a TRAP transporter from Desulfovibrio alaskensis, thought to import isethionate. Preliminary data has indicated the membrane components of the TRAP transporter have superior overexpression and stability compared to all other homologues screened. These properties will be more suitable for downstream experiments, when compared to other TRAP transporters that have been previously characterized. I have already solved the crystal structure of the substrate binding protein and revealed the interaction with isethionate. Further work will include the development of nanobodies for cryo-EM experiments on the membrane components, as well as small angle neutron scattering and proteoliposome assays to decipher the complete mechanism by which isethionate is imported. This will be the first structural and functional characterization of a non-sialic acid TRAP transporter. As these transporters are only found in bacteria and archaea, and share little homology to human transporters, they are an ideal family of transporters to target for antibiotic development. Hence the results of this project will underpin the future development of novel antibiotics.