Enabling Genetic Code Expansion and Peptide Macrocyclization in mRNA Display via a Promiscuous Orthogonal Aminoacyl-tRNA Synthetase
Albert Bowers
Associate Professor, UNC Chapel Hill
mRNA display is revolutionizing peptide drug discovery through its ability to quickly identify potent peptide binders of therapeutic protein targets. Methods to expand the chemical diversity of display libraries are continually needed to increase the likelihood of identifying clinically relevant peptide ligands. Orthogonal aminoacyl-tRNA synthetases (ORSs) have proven utility in cellular genetic code expansion, but are relatively underexplored for in vitro translation (IVT) and mRNA display. Herein, we demonstrate that the promiscuous ORS p-CNF-RS can incorporate noncanonical amino acids at amber codons in IVT, including the novel substrate p-cyanopyridylalanine (p-CNpyrA), to enable a pyridine–thiazoline (pyr–thn) macrocyclization in mRNA display. Pyr–thn-based selections against the deubiquitinase USP15 yielded a potent macrocyclic binder that exhibits good selectivity for USP15 and close homologues over other ubiquitin-specific proteases (USPs). Overall, this work exemplifies how promiscuous ORSs can both expand side chain diversity and provide structural novelty in mRNA display libraries through a heterocycle forming macrocyclization.
Born and raised in Trenton, NJ, Dr. Bowers obtained his BA from the University of Chicago and went on to complete his Ph.D. in organic chemistry in 2007, at the University of Illinois at Chicago, under the guidance of Prof. David Crich. He was an NIH postdoctoral fellow in the labs of Professor Robert M. Williams at Colorado State University and Professor Christopher T. Walsh at Harvard Medical School, where he studied the chemistry and biosynthesis of peptide-based natural products. In 2012, he joined the faculty at the University of North Carolina at Chapel Hill in 2012. He is jointly appointed in the School of Pharmacy and Department of Chemistry. The labs research focuses on biochemical and biosynthetic methods for the discovery of new peptide therapeutics, including mRNA display and macrocycles. For this work, Dr. Bowers has won a number of awards including the Beckman Young Investigator Award and Young Investigator Award of the Boulder Peptide Society.