Methods, cells and systems for incorporating non-canonical amino acids into proteins
AssigneeWaterstone Pharmaceuticals
InventorsYing Huang, Wenshe Liu, Ying Huang
Life of this patent
Estimating the term…
Abstract
The present disclosure pertains to methods of incorporating one or more non-canonical amino acids into a protein during a translation of the protein in bacterial cells. The present disclosure also pertains to methods of incorporating one or more non-canonical amino acids into a protein during an in vitro translation of the protein. In additional embodiments, the present disclosure pertains to isolated bacterial cells and in vitro translation systems (e.g., cell-free extract systems) for incorporating one or more non-canonical amino acids into a protein during a translation of the protein.
Claims37
A method of incorporating a non-canonical amino acid into a protein during a translation of said protein in bacterial cells, said method comprising:
- providing a modified ribosome in said bacterial cells, wherein said modified ribosome comprises a truncated L11 protein;
- providing a nucleotide sequence in said bacterial cells,
- wherein said nucleotide sequence encodes said protein, and
- wherein said nucleotide sequence comprises an introduced stop codon at a site where said incorporation of said non-canonical amino acid into said protein is desired;
- providing a suppressor transfer RNA in said bacterial cells,
- wherein said suppressor transfer RNA is acylated with said non-canonical amino acid, and
- wherein the anti-codon of said suppressor transfer RNA recognizes said introduced stop codon in said nucleotide sequence;
- and
- wherein a translation of said transcript by said modified ribosome and said suppressor transfer RNA incorporates said non-canonical amino acid into said translated protein.
A method of incorporating a non-canonical amino acid into a protein during an in vitro translation of said protein, said method comprising:
- providing a modified ribosome, wherein said modified ribosome comprises a truncated L11 protein;
- providing a nucleotide sequence,
- wherein said nucleotide sequence encodes said protein, and
- wherein said nucleotide sequence comprises an introduced stop codon at a site where said incorporation of said non-canonical amino acid into said protein is desired;
- providing a suppressor transfer RNA,
- wherein said suppressor transfer RNA is acylated with said non-canonical amino acid, and
- wherein the anticodon of said suppressor transfer RNA recognizes said introduced stop codon in said nucleotide sequence;
- and
- wherein a translation of said transcript by said modified ribosome and said suppressor transfer RNA incorporates said non-canonical amino acid into said translated protein.
An in vitro translation system for incorporating a non-canonical amino acid into a protein during a translation of said protein in said system, said system comprising:
- a modified ribosome, wherein said modified ribosome comprises a truncated L11 protein;
- a nucleotide sequence,
- wherein said nucleotide sequence encodes said protein, and
- wherein said nucleotide sequence comprises an introduced stop codon at a site where said incorporation of said non-canonical amino acid into said protein is desired;
- a suppressor transfer RNA,
- wherein said suppressor transfer RNA is acylated with said non-canonical amino acid, and
- wherein the anti-codon of said suppressor transfer RNA recognizes said introduced stop codon in said nucleotide sequence;
- and
- wherein said translation of said transcript by said modified ribosome and said suppressor transfer RNA incorporates said non-canonical amino acid into said translated protein.
An isolated bacterial cell for incorporating a non-canonical amino acid into a protein during a translation of said protein in said bacterial cell, said bacterial cell comprising:
- a modified ribosome, wherein said modified ribosome comprises a truncated L11 protein;
- a nucleotide sequence,
- wherein said nucleotide sequence encodes said protein, and
- wherein said nucleotide sequence comprises an introduced stop codon at a site where said incorporation of said non-canonical amino acid into said protein is desired,
- a suppressor transfer RNA,
- wherein said suppressor transfer RNA is acylated with said non-canonical amino acid, and
- wherein the anti-codon of said suppressor transfer RNA recognizes said introduced stop codon in said nucleotide sequence;
- and
- wherein said translation of said transcript by said modified ribosome and said suppressor transfer RNA incorporates said non-canonical amino acid into said translated protein.
Dependencies are inferred from the claim text.