Expanding the Scope of Sortase‐Mediated Ligations by Using Sortase Homologues
2017; Wiley; Volume: 19; Issue: 2 Linguagem: Inglês
10.1002/cbic.201700517
ISSN1439-7633
AutoresKeyvan D. Nikghalb, Nicholas Horvath, Jesse Prelesnik, Orion GB Banks, Pavel A. Filipov, R. David Row, Travis J. Roark, John M. Antos,
Tópico(s)Biochemical and Structural Characterization
ResumoSortase-catalyzed transacylation reactions are widely used for the construction of non-natural protein derivatives. However, the most commonly used enzyme for these strategies (sortase A from Staphylococcus aureus) is limited by its narrow substrate scope. To expand the range of substrates compatible with sortase-mediated reactions, we characterized the in vitro substrate preferences of eight sortase A homologues. From these studies, we identified sortase A enzymes that recognize multiple substrates that are unreactive toward sortase A from S. aureus. We further exploited the ability of sortase A from Streptococcus pneumoniae to recognize an LPATS substrate to perform a site-specific modification of the N-terminal serine residue in the naturally occurring antimicrobial peptide DCD-1L. Finally, we unexpectedly observed that certain substrates (LPATXG, X=Nle, Leu, Phe, Tyr) were susceptible to transacylation at alternative sites within the substrate motif, and sortase A from S. pneumoniae was capable of forming oligomers. Overall, this work provides a foundation for the further development of sortase enzymes for use in protein modification.
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