Selective Late-Stage Functionalization of Tryptophan-Containing Peptides To Facilitate Bioorthogonal Tetrazine Ligation

Abstract

Site-selective modification of complex peptides and the functionalization of their C−H bonds hold great promise for expanding their use in therapeutics and biomedical research. Herein, we leverage the power of late-stage chemoenzymatic catalysis using an indole prenyltransferase (IPT) enzyme and alkyl diphosphates to specifically modify the indole ring of tryptophan in clinically relevant peptides. Furthermore, the installed handle enables bioorthogonal click chemistry through an inverse electrondemand Diels−Alder (IEDDA) reaction with a biotin-conjugated tetrazine probe.

Chemical Synthesis of Human Proteoforms and Application in Biomedicine

“…The unnatural amino acids can carry modifications such as acylation and phosphorylation, − while more complex PTMs (e.g., glycosylation, ubiquitination) remain difficult to incorporate due to challenge in the engineering of ribosomal aminoacyl-tRNA synthetases (Figure a). Second, bio-orthogonal functionalization techniques enable the preparation of site-selectively modified proteins at specific residues such as Cys. − Lys, − Met, − and Trp − of expressed protein (Figure b). PTM mimics are usually produced through this technology, which, however, may occasionally result in unforeseen biological consequences or ambiguity in interpreting experimental findings .…”