A pyrrolysine-tRNA synthetase for site-specific insertion of NAEK
By using a combination of pyrrolidone-lysine-tRNA synthetase derived from *Methanococcus hortensia* and orthogonal tRNA, the problem of limited insertion range of non-natural amino acids in *Escherichia coli* was solved, achieving efficient site-directed insertion and a significant increase in the yield of the target protein.
CN122256274APending Publication Date: 2026-06-23NINGBO INST OF MARINE MEDICINE PEKING UNIV
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- NINGBO INST OF MARINE MEDICINE PEKING UNIV
- Filing Date
- 2026-03-19
- Publication Date
- 2026-06-23
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Figure CN122256274A_ABST
Abstract
The application discloses a pyrrolysine-tRNA synthetase for site-specific insertion of NAEK, and belongs to the technical field of enzyme engineering. The pyrrolysine-tRNA synthetase can be expressed heterogeneously in Escherichia coli, and has excellent enzymatic properties, high enzyme activity, high expression amount and good solubility. In a high-density culture medium, the yield of IL2(Y45) of Mh PylRS for site-specific insertion of NAEK at a TAG termination codon is increased by 94% compared with that of the currently commonly used Mb PylRS, and is increased by 23% compared with that of Mm PylRS; the yield of IL2(Y45) of Mh PylRS for site-specific insertion of NAEK at a UGA termination codon is increased by 96% compared with that of the currently commonly used Mb PylRS, and is increased by 28% compared with that of Mm PylRS. The enzyme for efficient insertion of NAEK not only improves the yield of the target protein, but also lays a foundation for subsequent simultaneous insertion of various unnatural amino acids.
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