Genetic Code Expansion of the Silkworm Bombyx mori Using a Pyrrolysyl-tRNA Synthetase/tRNA^Pyl Pair

The domesticated silkworm Bombyx mori, an essential industrial animal for silk production, has attracted attention as a host for protein production due to its remarkable protein synthesis capability. Here, we applied genetic code expansion (GCE) using a versatile pyrrolysyl-tRNA synthetase (PylRS)/tRNA^Pyl pair from Methanosarcina mazei to B. mori; GCE enables synthetic amino acid incorporation into proteins to give them non-natural functions. Transgenic B. mori lines expressing M. mazei PylRS and its cognate tRNA^Pyl were generated and cross-mated to obtain their F₁ hybrid. Orally administering a click-compatible synthetic amino acid, trans-cyclooctene-lysine (TCO-Lys), to the F₁ hybrid has led to the production of silk fiber incorporated with TCO-Lys. TCO-Lys incorporation in silk fiber was verified by selective labeling of the TCO group by click chemistry. The developed system is available for large-scale protein production with a wide variety of synthetic amino acids.