Abstract
Over 20 years ago, the pyrrolysine encoding translation system was discovered in specific archaea. Our Review provides an overview of how the once obscure pyrrolysyl-tRNA synthetase (PylRS) tRNA pair, originally responsible for accurately translating enzymes crucial in methanogenic metabolic pathways, laid the foundation for the burgeoning field of genetic code expansion. Our primary focus is the discussion of how to successfully engineer the PylRS to recognize new substrates and exhibit higher in vivo activity. We have compiled a comprehensive list of ncAAs incorporable with the PylRS system. Additionally, we also summarize recent successful applications of the PylRS system in creating innovative therapeutic solutions, such as new antibody-drug conjugates, advancements in vaccine modalities, and the potential production of new antimicrobials.
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