Rapid Discovery Platform for Orthogonal tRNA Synthetase/tRNA Pairs Expands the Non-Canonical Amino Acid Toolbox
Cervettini, D., Tang, S., Fried, S.D., Willis, J.C.W., Funke, L.F.H., Colwell, L.J., Chin, J.W.
Nature Biotechnology 38, 989 (2020)
Published in Nature Biotechnology in 2020, this paper describes a platform for the rapid discovery and evolution of orthogonal aminoacyl-tRNA synthetase/tRNA pairs, the molecular components that determine which non-canonical amino acids can be genetically encoded.
Before this work, developing a new orthogonal synthetase/tRNA pair for a specific ncAA was a months-long process. Each pair had to be individually engineered: the synthetase needed to recognise the target ncAA while ignoring all 20 canonical amino acids, and the tRNA needed to function in translation while avoiding recognition by the cell's native synthetases. The selection and evolution methods available were effective but slow, limiting the rate at which new ncAAs could be added to the genetic code expansion toolbox.
This platform accelerated the process by combining improved library design, more efficient selection methods, and machine learning-guided approaches to identify functional synthetase/tRNA pairs from large sequence spaces. Rather than engineering each new pair from scratch, the platform enables systematic discovery across diverse ncAA chemistries.
The practical consequence is a faster path from identifying a desirable ncAA chemistry to having the molecular machinery needed to incorporate it into proteins via fermentation. For Constructive Bio's therapeutic programmes, this means that when a drug development programme requires a specific modification, such as a particular conjugation handle, a protease-resistant backbone modification, or a lipidation chemistry, the synthetase/tRNA pair needed to encode that modification can be identified and validated more rapidly.
This platform, together with the automated orthogonal tRNA generation published in 2024 and the orthogonal replication system for accelerated evolution published in the same year, forms the engineering infrastructure that continuously expands the ncAA repertoire available through BioForge manufacturing.
Why it matters
The range of therapeutic applications depends on how many different ncAAs can be incorporated. This rapid discovery platform means Constructive Bio can quickly develop the molecular machinery for new ncAA types requested by pharmaceutical partners, rather than spending years engineering each one. It is the discovery engine that feeds the BioForge manufacturing platform.
Explore our platform
See how this research translates into next-generation peptide therapeutics.