Expanding the Scope of Sortase‐Mediated Ligations by Using Sortase Homologues |
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Authors: | Keyvan D Nikghalb Nicholas M Horvath Jesse L Prelesnik Orion G B Banks Pavel A Filipov R David Row Travis J Roark Prof John M Antos |
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Affiliation: | Department of Chemistry, Western Washington University, Bellingham, WA, USA |
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Abstract: | Sortase‐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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Keywords: | enzymes peptides protein modifications sortase transacylation |
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