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Structural and functional characterization of SplA, an exclusively specific protease of Staphylococcus aureus

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  • Justyna Stec-Niemczyk, Denmark
  • Katarzyna Pustelny, Denmark
  • Magdalena Kisielewska, Denmark
  • Michal Bista, Denmark
  • Kevin Boulware, Denmark
  • Henning Stennicke, Denmark
  • Ida Thøgersen
  • Patrick Daugherty, Denmark
  • Jan Enghild
  • Grzegorz Popowicz, Denmark
  • Adam Dubin, Denmark
  • Jan Potempa, Denmark
  • Grzegorz Dubin, Denmark
Staphylococcus aureus is a dangerous human pathogen which antibiotic resistance is steadily increasing and no efficient vaccine is as yet available. This serious threat drives extensive studies on staphylococcal physiology and pathogenicity pathways, especially virulence factors. Serine protease like proteins (Spl) encoded by an operon containing up to six genes are a good example of poorly characterized secreted proteins likely involved in virulence. Here we describe an efficient heterologous expression system for SplA and detailed biochemical and structural characterization of the recombinant SplA protease. The enzyme shares a significant sequence homology to V8 protease and epidermolytic toxins which are well documented staphylococcal virulence factors. SplA has a very narrow substrate specificity apparently imposed by the precise recognition of three amino acid residues positioned N-terminal to the hydrolyzed peptide bond. To explain determinants of this extended specificity we resolve the crystal structure of SplA and define consensus model of substrate binding. Furthermore we demonstrate that artificial N-terminal elongation of mature SplA mimicking a naturally present signal peptide abolishes enzymatic activity. The likely physiological role of the process is discussed. Of interest, even though precise N-terminal trimming is a common regulatory mechanism among S1 family enzymes, the crystal structure of SplA reveals novel, significantly different mechanistic details.
Original languageEnglish
JournalBiochemical Journal
ISSN0264-6021
DOIs
Publication statusPublished - 2009

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