1. Characterization of the housekeeping sortase from the human pathogen Propionibacterium acnes: first investigation of a class F sortase
- Author
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Roger R. Beerli, Salvatore Di Girolamo, Ulf Grawunder, Tamara Hell, Chasper Puorger, Mara Castiglione, Manfred Briendl, Georg Lipps, Rémy Gebleux, and Maren Vogel
- Subjects
chemistry.chemical_classification ,biology ,Cell Biology ,Protein engineering ,medicine.disease_cause ,biology.organism_classification ,Biochemistry ,Propionibacterium acnes ,Enzyme ,chemistry ,Recognition sequence ,Staphylococcus aureus ,Sortase ,Sortase A ,medicine ,Molecular Biology ,Bacteria - Abstract
Sortase enzymes play an important role in Gram-positive bacteria. They are responsible for the covalent attachment of proteins to the surface of the bacteria and perform this task via a highly sequence-specific transpeptidation reaction. Since these immobilized proteins are often involved in pathogenicity of Gram-positive bacteria, characterization of this type of enzyme is also of medical relevance. Different classes of sortases (A–F) have been found, which recognize characteristic recognition sequences present in substrate proteins. Up to date, sortase A from Staphylococcus aureus, a housekeeping class A sortase, is the most thoroughly studied representative of the sortase family of enzymes. Here we report the in-depth characterization of the class F sortase from Propionibacterium acnes, a class of sortases that has not been investigated before. As Sortase F is the only transpeptidase found in the P. acnes genome, it is the housekeeping sortase of this organism. Sortase F from P. acnes shows a behavior similar to sortases from class A in terms of pH dependence, recognition sequence and catalytic activity; furthermore, its activity is independent of bivalent ions, which contrasts to sortase A from S. aureus. We demonstrate that sortase F is useful for protein engineering applications, by producing a site-specifically conjugated homogenous antibody–drug conjugate with a potency similar to that of a conjugate prepared with sortase A. Thus, the detailed characterization presented here will not only enable the development of anti-virulence agents targeting P. acnes but also provides a powerful alternative to sortase A for protein engineering applications.
- Published
- 2019
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