Unnatural reactive amino acid genetic code additions

Patent metadata

Field	Value
Publication number	US-9797908-B2
Application number	US-201414338113-A
Country	US
Kind code	B2
Filing date	Jul 22, 2014
Priority date	Jun 18, 2003
Publication date	Oct 24, 2017
Grant date	Oct 24, 2017

Abstract

This invention provides compositions and methods for producing translational components that expand the number of genetically encoded amino acids in eukaryotic cells. The components include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, orthogonal pairs of tRNAs/synthetases and unnatural amino acids. Proteins and methods of producing proteins with unnatural amino acids in eukaryotic cells are also provided.

First claim

What is claimed is:

1. A composition comprising a protein, antibody, or antibody fragment, wherein the protein, antibody or antibody fragment comprises at least one unnatural amino acid comprising a first reactive group and at least one post-translational modification, wherein the at least one post-translational modification comprises attachment of a molecule comprising a second reactive group by a cycloaddition reaction to the at least one unnatural amino acid comprising the first reactive group;
wherein the unnatural amino acid comprising the first reactive group was incorporated into the protein, antibody or antibody fragment using an orthogonal translation system.

2. The composition of claim 1 , wherein the first reactive group is an alkynyl or azido moiety and the second reactive group is an azido or alkynyl moiety.

3. The composition of claim 2 , wherein the first reactive group is the alkynyl moiety and the second reactive group is the azido moiety.

4. The composition of claim 2 , wherein the first reactive group is the azido moiety and the second reactive group is the alkynyl moiety.

5. The composition of claim 1 , wherein the at least one post-translational modification is made in vivo in a eukaryotic cell.

6. The composition of claim 1 , further comprising a eukaryotic cell containing the protein, antibody, or antibody fragment.

7. The composition of claim 1 , wherein the unnatural amino acid is a p-azido-L-phenylalanine or a p-propargyloxyphenylalanine.

8. The composition of claim 1 , wherein the molecule is a dye, a polymer, a derivative of polyethylene glycol, a photocrosslinker, a cytotoxic compound, an affinity label, a derivative of biotin, a resin, a second protein or polypeptide, a metal chelator, a cofactor, a fatty acid, a carbohydrate, or a polynucleotide.

9. The composition of claim 1 , wherein the at least one unnatural amino acid was incorporated into the protein, antibody, or antibody fragment using an orthogonal synthetase comprising a polypeptide sequence selected from the group consisting of any one of SEQ ID NOs.: 48 to 63.

Assignees

• Scripps Research Inst

Inventors

• Deiters Alexander
• Cropp T Ashton
• Chin Jason W
• Anderson J Christopher
• Schultz Peter G

Classifications

• A61P31/18

  for HIV · CPC title

• A61P3/10

  for hyperglycaemia, e.g. antidiabetics · CPC title

• A61P35/00

  Antineoplastic agents · CPC title

• C07C229/36

  with at least one amino group and one carboxyl group bound to the same carbon atom of the carbon skeleton · CPC title

• G01N33/583 Primary

  with non-fluorescent dye label · CPC title