Oral Presentation 49th Lorne Conference on Protein Structure and Function 2024

Extracellular loops of the β-barrel domain mediate rapid folding and function of self-associating autotransporters (#12)

Xiaojun Yuan 1 , Matthew D Johnson 1 , Jiansi Long 1 , Jing Zhang 1 , Alvin W Lo 2 3 , Jason J Paxman 4 , Minh Duy Phan 2 3 , Begoña Heras 4 , Mark A Schembri 2 3 , Ian R Henderson 2 5 , Denisse L Leyton 1
  1. Research School of Biology , Australian National University, Canberra , ACT, Australia
  2. School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Queensland, Australia
  3. Australian Infectious Disease Research Centre, The University of Queensland, Brisbane, Queensland, Australia
  4. Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, VIC, Australia
  5. Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland, Australia

Bacterial aggregation is a phenotype associated with disease pathogenesis. Aggregate formation enhances biofilm development, host colonization, and resistance to antibiotics and host defenses. Antigen 43 (Ag43) is a surface-located autotransporter produced by pathogenic Escherichia coli that mediates cell aggregation in biofilms. Here, two Ag43 passenger domains from neighboring bacterial cells fold into elongated β-helices and associate in a head-to-tail manner while being anchored to the cell surface by their outer membrane-embedded β-barrel domains. In this study, we use mutational analysis on Ag43 to show that the β-hairpin structure of the fourth and fifth extracellular loops of the β-barrel domain have a crucial role for passenger domain folding and subsequent formation of bacterial aggregates. Mutagenesis of the homologous autotransporter, adhesin involved in diffuse adherence (AIDA-I), suggests that the requirement of these long extracellular loops for passenger folding and biological function is conserved among self-associating autotransporters. This work provides mechanistic insight into the role of the autotransporter β-barrel domain as a folding vector that mediates rapid folding of the passenger domain into the β-helix that enables bacterial interactions during infection.