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Enzymatic C-to-C Protein Ligation.

ABSTRACT: Transpeptidase-catalyzed protein and peptide modifications have been widely utilized for generating conjugates of interest for biological investigation or therapeutic applications. However, all known transpeptidases are constrained to ligating in the N-to-C orientation, limiting the scope of attainable products. Here, we report that an engineered asparaginyl ligase accepts diverse incoming nucleophile substrate mimetics, particularly when a means of selectively quenching the reactivity of byproducts released from the recognition sequence is employed. In addition to directly catalyzing formation of l-/d- or α-/β-amino acid junctions, we find C-terminal Leu-ethylenediamine (Leu-Eda) motifs to be bona fide mimetics of native N-terminal Gly-Leu sequences. Appending a C-terminal Leu-Eda to synthetic peptides or, via an intein-splicing approach, to recombinant proteins enables direct transpeptidase-catalyzed C-to-C ligations. This work significantly expands the synthetic scope of enzyme-catalyzed protein transpeptidation reactions.

SUBMITTER: Rehm FBH

PROVIDER: S-EPMC9303898 | biostudies-literature |

REPOSITORIES: biostudies-literature

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