# US 2010/0167347 A1 · Compositions of Aminoacyl-tRNA Synthetase and Uses Thereof

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- Number: US 2010/0167347 A1
- Document ID: US20100167347A1
- Status: Published application
- Kind: A1
- Application number: 12/724,365
- Filed: 2010-03-15
- Published: 2010-07-01
- Earliest claimed: 2004-12-22
- Assignees: Ambrx, Inc.
- Inventors: Andrew Paulsel; Ho Sung Cho
- Web page: https://patentgrep.com/patent/US20100167347A1

## CPC

- C12N 9/93: {Ligases (6)}
- C12P 21/02: having a known sequence of two or more amino acids, e.g. glutathione
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)}

## Abstract

Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNA's, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNA's/synthetases are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins using these orthogonal pairs.

## Claims

### Claim at position 1

1-18. (canceled)

### Claim 19

A cell comprising a translation system, wherein the translation system comprises an aminoacyl tRNA synthetase (RS) nucleic acid sequence selected from the group consisting of SEQ ID NO: 17, and a complementary polynucleotide sequence of SEQ ID NO: 17.

### Claim 20 (depends on 19)

The cell of claim 19, wherein the cell is a eukaryotic cell.

### Claim 21 (depends on 20)

The cell of claim 20, wherein the eukaryotic cell is a yeast cell.

### Claim 22 (depends on 20)

The cell of claim 20, wherein the eukaryotic cell is a fungal cell.

### Claim 23 (depends on 20)

The cell of claim 20, wherein the eukaryotic cell is a mammalian cell.

### Claim 24 (depends on 20)

The cell of claim 20, wherein the eukaryotic cell is an insect cell.

### Claim 25 (depends on 20)

The cell of claim 20, wherein the eukaryotic cell is a plant cell.

### Claim 26 (depends on 19)

The cell of claim 19, wherein the cell is a non-eukaryotic cell.

### Claim 27 (depends on 26)

The cell of claim 26, wherein the non-eukaryotic cell is an E. coli cell.

### Claim 28 (depends on 19)

The cell of claim 19, further comprising a tRNA that is aminoacylated by the RS and polynucleotide encoding a polypeptide of interest, wherein the polynucleotide comprises a selector codon that is recognized by the tRNA.

### Claim 29 (depends on 28)

The cell of claim 28, wherein said polypeptide of interest is human growth hormone.

### Claim 30 (depends on 19)

The cell of claim 19, wherein the cell is an E. coli.

### Claim at position 14

31-35. (canceled)

### Claim 36

A vector comprising a polynucleotide encoding an RS having a polynucleotide sequence encoding the amino acid sequence of SEQ ID NO: 17, and a complementary polynucleotide sequence thereof.

### Claim 37 (depends on 36)

The vector of claim 36, wherein the vector comprises a plasmid, a cosmid, a phage, or a virus.

### Claim 38 (depends on 36)

The vector of claim 36, wherein the vector is an expression vector.

### Claim 39 (depends on 36)

A cell comprising the vector of claim 36.

### Claim 40

A method of producing a polypeptide in a cell with a selected amino acid at a specified position, the method comprising:
growing, in an appropriate medium, the cell, where the cell comprises a nucleic acid that comprises at least one selector codon and encodes a polypeptide; and,
providing the selected amino acid;
wherein the cell further comprises:
an orthogonal RS (O-RS) that functions in the cell wherein said RS has an amino acid sequence selected from the group consisting of SEQ ID NO: 17, and a complementary polynucleotide sequence thereof; and,
an orthogonal tRNA (O-tRNA), that recognizes the selector codon wherein said O-RS aminoacylates the O-tRNA with the selected amino acid.

### Claim 41 (depends on 40)

The method of claim 40 wherein said selected amino acid is para-acetyl phenylalanine.

### Claim at position 21

42-43. (canceled)

### Claim 44 (depends on 40)

The method of claim 40 wherein said polypeptide is human growth hormone.

### Claim 45

A polynucleotide having a polynucleotide sequence encoding the amino acid sequence of SEQ ID NO: 17, and a complementary polynucleotide sequence thereof.