Sortase mediated multi‐fragment assemblies by ligation site switching
نویسندگان
چکیده
Die Sortase-vermittelte Ligation (Engl. sortase-mediated ligation, SML) ist ein effektives Werkzeug der Proteinchemie, welches die von Peptiden mit LPxTG-Sortiermotiven und N-terminalen Glycin-Nukleophilen ermöglicht. Kopplung mehrerer Fragmente durch SML wird den erneuten Einbau eines Sortiermotivs in das Produkt verhindert. Hier beschreiben wir Multi-Fragment-SML auf Grundlage schaltbaren Sortase-Substraten. Substitution des Leu-Restes disulfidhaltiges Cys(StBu) führt zu aktiven Sortiermotiven, Reduktion inaktiviert werden können. In Kombination einem photo-geschützten N-Gly-Nukleophil wiederholte Zyklen Ligationsstellenschaltung Durchführbarkeit dieser Herangehensweise wurde eine Proof-of-Concept Vier-Fragment-Ligation, Fertigung Peptidsonden für bivalente chromatinbindende Proteine sowie Oligomerisierung Peptidantigenen nachgewiesen. Biochemische immunologische Assays zeigten Funktionalität Sonden, was sie vielversprechenden Werkzeugen Immunologie Chromatinbiochemie macht.
منابع مشابه
Sortase-mediated protein ligation: a new method for protein engineering.
Sortase (SrtA), a transpeptidase from Staphylococcus aureus, catalyzes a cell-wall sorting reaction at an LPXTG motif by cleaving between threonine and glycine and subsequently joining the carboxyl group of threonine to an amino group of pentaglycine on the cell wall peptidoglycan. We have applied this transpeptidyl activity of sortase to in vitro protein ligation. We found that in the presence...
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ژورنال
عنوان ژورنال: Angewandte Chemie
سال: 2021
ISSN: ['1521-3773', '1433-7851', '0570-0833']
DOI: https://doi.org/10.1002/ange.202109032