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

The identification and characterisation of an unusual disulfide bond forming protein from Salmonella Typhimurium (#422)

Pramod Subedi 1 2 , Jason Paxman 1 , Tony Wang 1 , Lilian Hor 1 , Makrina Totsika 3 , Jennifer Martin 4 , Begoña Heras 1
  1. Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria 3086, Australia, Thornburry, VIC, Australia
  2. School of Chemistry and Bio21, Molecular Science and Biotechnology Institute, , University of Melbourne, Parkville, Victoria 3010, Australia
  3. Institute of Health and Biomedical Innovation, , School of Biomedical Sciences, Queensland University of Technology,, Brisbane,, Queensland,, Australia
  4. Griffith Institute for Drug Discovery , Nathan, QLD , Australia

Bacteria use folding enzymes to produce functional virulence factors. These foldases include the Dsb family of proteins, which catalyze a key step in the protein-folding pathway, the introduction of disulfide bonds. The Dsb oxidative system, which includes an oxidative DsbA/DsbB pathway and an isomerase DsbC/DsbD pathway, is present in numerous bacterial species. Conventionally, Dsb proteins have specific redox functions with monomeric and dimeric Dsbs exclusively catalyzing thiol oxidation and disulfide isomerization, respectively. This contrasts with the eukaryotic disulfide forming machinery where the modular thioredoxin protein PDI mediates thiol oxidation and disulfide reshuffling.

In this study, we identified and structurally and biochemically characterized a novel Dsb-like protein from Salmonella enterica termed BcfH and defined its role in virulence.

Encoded by a highly conserved bcf (bovine colonization factor) fimbrial operon, the Dsb-like enzyme BcfH forms a trimeric structure, exceptionally uncommon among the large and evolutionary conserved thioredoxin superfamily. BcfH also displays unusual catalytic redox centers, including an unwound α-helix holding the redox active site and a trans proline instead of the conserved cis proline active site loop. Remarkably, BcfH displays both thiol oxidase and disulfide isomerase activities contributing to Salmonella fimbrial biogenesis. Typically, oligomerization of bacterial Dsb proteins modulates their redox function, with monomeric and dimeric Dsbs mediating thiol oxidation and disulfide isomerization, respectively. The present study demonstrates a further structural and functional malleability in the thioredoxin-fold protein family. BcfH trimeric architecture and unconventional catalytic sites permit multiple redox functions emulating in bacteria the eukaryotic protein disulfide isomerase dual oxido-reductase activity.

The outcomes of this research will also establish how bacterial pathogens utilise multipleDsbhomologuesto generate virulence proteins, which is essential for understanding how bacteria cause disease.