Site-Specific Protein Labeling and Generation of Defined Ubiquitin-Protein Conjugates Using an Asparaginyl Endopeptidase.

ABSTRACT: Asparaginyl endopeptidases (AEPs) have recently been widely utilized for peptide and protein modification. Labeling is however restricted to protein termini, severely limiting flexibility and scope in creating diverse conjugates as needed for therapeutic and diagnostic applications. Here, we use genetic code expansion to site-specifically modify target proteins with an isopeptide-linked glycylglycine moiety that serves as an acceptor nucleophile in AEP-mediated transpeptidation with various probes containing a tripeptidic recognition motif. Our approach allows simple and flexible labeling of recombinant proteins at any internal site and leaves a minimal, entirely peptidic footprint (NGG) in the conjugation product. We show site-specific labeling of diverse target proteins with various biop

SUBMITTER: Fottner M 

PROVIDER: S-EPMC9335880 | biostudies-literature | 2022 Jul

REPOSITORIES: biostudies-literature