Oral Presentation The 35th Biennial Conference of the Society of Crystallographers in Australia and New Zealand 2024 (Crystal 35)

Unravelling Autotransporter Mechanisms: A Path to New Anti-Virulence Therapies (109116)

Jason Paxman 1 , Lilian Hor 1 , Akila Pilapitiya 1 , Mark Schembri 2 , Begoña Heras 1
  1. Department of Biochemistry and Chemistry, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, VIC, Australia
  2. School of Chemistry and Molecular Biosciences , The University of Queensland, Brisbane, QLD 4072, Australia

Antimicrobial Resistance (AMR) is one of the most pressing global health threats of our time, demanding immediate and innovative solutions. Our research focuses on the virulence proteins that bacteria use as their molecular arsenal to cause disease. By combining structural biology  with advanced molecular and biochemical techniques, we are uncovering the intricate mechanisms behind key bacterial virulence factors.

 At the heart of our work lies the Type V secretion system, responsible for secreting autotransporters—the largest group of secreted proteins in Gram-negative bacteria [1]. These proteins play crucial roles in pathogenesis, including promoting bacterial aggregation and biofilm formation, along with facilitating adherence to host cells, and causing cytotoxicity.

Our multidisciplinary approach that combines X-ray crystallography, biophysical techniques, immunoassays, and cellular assays, has allowed us to elucidate the structure and function of diverse autotransporter proteins, including adhesins and cytotoxins, revealing how they facilitate their pathogenic functions [2-5]. For example, our findings indicate that different autotransporter adhesins promote bacterial aggregation and biofilm formation through a conserved self-association mechanism, facilitating host colonization and bacterial persistence in various environmental niches [2-3]. We are also uncovering, in atomic detail, how autotransporter toxins such as the proteases Ssp and EspC invade host cells to cause cytotoxic damage.

This understanding is enabling us to develop antimicrobials that "disarm rather than kill bacteria," a novel strategy aimed at minimising resistance development. We are actively creating autotransporter inhibitors, including our patented biofilm blocker [6], which can serve as stand-alone antibiofilm therapy or enhance the efficacy of failing antibiotics. We are also developing molecules that block autotransporter toxins laying the groundwork for future anti-toxin drugs.

Overall, our research not only provides a deeper understanding of bacterial pathogenesis but also opens new avenues for combating AMR.

  1. Clarke KR, Hor L, Pilapitiya A, Luirink J, Paxman JJ and Heras B. Phylogenetic Classification and Functional Review of Autotransporters. 2022.Frontiers in Immunology. 13, 921272
  2. Heras B, Totsika M, Peters KM, Paxman JJ, Gee C, Jarrott R, Perugini MA, Whitten AE, Schembri MA. The antigen 43 structure reveals a molecular Velcro-like mechanism of autotransporter-mediated bacterial clumping. Proc Natl Acad Sci USA. 2014; 111, 457-462.
  3. Vo JL, Martínez Ortiz GC, Totsika M, Lo A, Hancock S, Whitten A, Hor L, Peters K, Ageorges V, Caccia N, Desvaux M, Schembri MA, Paxman JJ and Heras B. Variation of Antigen 43 self-association modulates bacterial compacting within aggregates and biofilms. npj Biofilms Microbiomes 8, 20 2022
  4. Paxman JJ, Lo A, Sullivan MJ, Panjikar S, Kuiper M, Whitten AW, Wang G, Luan CH, Moriel DG, Tan L, Peters KM, Phan MD, Gee CL, Ulett GC, Schembri MA , Heras B. Unique structural features of a bacterial autotransporter adhesin suggest mechanisms for interaction with host macromolecules. Nat Commun. 2019 Apr 29;10(1):1967.
  5. Hor L, Pilapitiya A, McKenna JA, Panjikar S, Anderson MA, Desvaux M, Paxman JJ, Heras B. Crystal structure of a subtilisin-like autotransporter passenger domain reveals insights into its cytotoxic function. 2023, Nat Commun. 14(1):1163.
  6. Heras, B., Paxman J.J., Schembri, M.A., Lo, A., PCT/AU2019/050893 (2019).