Proteolytic Profiling of Streptococcal Pyrogenic Exotoxin B (SpeB) by Complementary HPLC-MS Approaches

Constantin Blöchl, Christoph Holzner, Michela Luciano, Renate Bauer, Jutta Horejs-Hoeck, Ulrich Eckhard, Hans Brandstetter, Christian G. Huber

Research output: Contribution to journalArticlepeer-review

Abstract

Streptococcal pyrogenic exotoxin B (SpeB) is a cysteine protease expressed during group A streptococcal infection that represents a major virulence factor. Although subject to several studies, its role during infection is still under debate, and its proteolytic properties remain insufficiently characterized. Here, we revisited this protease through a set of complementary approaches relying on state of-the-art HPLC-MS methods. After conceiving an efficient protocol to recombinantly express SpeB, the zymogen of the protease and its activation were characterized. Employing proteome-derived peptide libraries, a strong preference for hydrophobic and aromatic residues at P2 alongside negatively charged amino acids at P3′ to P6′ was revealed. To identify relevant in vivo substrates, native proteins were obtained from monocytic secretome and plasma to assess their cleavage under physiological conditions. Besides corroborating our findings concerning specificity, more than 200 cleaved proteins were identified, including proteins of the extracellular matrix, proteins of the immune system, and proteins involved in inflammation. Finally, the cleavage of IgG subclasses was studied in detail. This study precisely depicts the proteolytic properties of SpeB and provides a library of potential host substrates, including their exact cleavage positions, as a valuable source for further research to unravel the role of SpeB during streptococcal infection.
Original languageEnglish
Article number412
JournalInternational Journal of Molecular Sciences
Volume23
Issue number1
DOIs
Publication statusPublished - 30 Dec 2021

Bibliographical note

Publisher Copyright:
© 2021 by the authors. Licensee MDPI, Basel, Switzerland.

Keywords

  • HUNTER
  • IgG subclasses
  • N-terminomics
  • Positional proteomics
  • Protease degradomics
  • SCP
  • Sequence logo
  • Sequence specificity
  • Streptococcal cysteine protease
  • Streptopain
  • sequence logo
  • sequence specificity
  • streptococcal cysteine protease
  • protease degradomics
  • positional proteomics
  • streptopain

Fields of Science and Technology Classification 2012

  • 104 Chemistry
  • 106 Biology

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