# US 10,370,776 B2 · Antibody like protein

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 10,370,776 B2
- Document ID: US10370776B2
- Status: Granted
- Kind: B2
- Application number: 14/497,320
- Filed: 2014-09-25
- Published: 2016-03-31
- Granted: 2019-08-06
- Earliest claimed: 2013-09-25
- Assignees: Idea Orchard, LLC
- Applicants: Michael Longo; Rajika Perera
- Inventors: Michael Longo; Rajika Perera
- Web page: https://patentgrep.com/patent/US10370776B2

## CPC

- C40B 40/10: Libraries containing peptides or polypeptides, or derivatives thereof
- C12N 15/102: {Mutagenizing nucleic acids}
- C12N 15/1031: {mutagenesis by gene assembly, e.g. assembly by oligonucleotide extension PCR}
- C12N 15/1037: {Screening libraries presented on the surface of microorganisms, e.g. phage display, E. coli display}
- C40B 10/00: Directed molecular evolution of macromolecules, e.g. RNA, DNA or proteins
- C40B 40/08: Libraries containing RNA or DNA which encodes proteins, e.g. gene libraries
- C07K 2319/20: containing a tag with affinity for a non-protein ligand
- C07K 2319/50: containing protease site
- C07K 2319/70: containing domain for protein-protein interaction
- C12N 15/1034: {Isolating an individual clone by screening libraries}
- C12N 15/1058: {Directional evolution of libraries, e.g. evolution of libraries is achieved by mutagenesis and screening or selection of mixed population of organisms}
- C40B: COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES (in silico combinatorial libraries of nucleic acids, proteins or peptides G16B35/00; in silico combinatorial chemistry G16C20/60)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)

## Abstract

A general method and recombinant nucleic acid sequences, by means which the method selects a recombinant protein containing an FHA domain for binding a target molecule from a library proteins with a high-throughput method of creating protein variations within the FHA domain in non-conserved or non-structural sequences of the FHA scaffold, and the library may also be in the form of a phagemid or phage library wherein the ALP nucleic acid sequence is inserted into a vector capable of allowing the vector and expressed ALP protein from being virally packaged, and the recombinant nucleic acid sequences which are randomly mutated at varying non-conserved or non-structural FHA domain sequences.

## Claims

### Claim 1

A protein having an FHA domain comprising:
a. a scaffold domain and loop domains derived from a wild-type FHA domain;
b. at least one mutated loop domain;
c. at least one mutated scaffold sequence between the loops in regions that are non-highly conserved among wild-type FHA domains; and
d. the protein having specific binding affinity to a target molecule that does not include interactions with phosphate moieties based on naturally occurring, or in vivo FHA domain recognition sites.

### Claim 2 (depends on 1)

The protein of claim 1 wherein the mutations include post-translational modifications to the amino acid residues.

### Claim 3 (depends on 1)

The protein of claim 1 wherein the mutations include a deletion, a replacement, or an addition to the amino acid residues.

### Claim 4 (depends on 1)

The protein of claim 1 may be used for preparative, analytical, or therapeutic methods that involve the binding of said protein to said target molecule.

### Claim 5 (depends on 1)

An isolated nucleic acid sequence encoding for said protein of claim 1.

### Claim 6 (depends on 5)

The isolated nucleic acid sequence of claim 5 that encodes said protein is transfected into a host cell.

### Claim 7 (depends on 5)

The isolated nucleic acid sequence of claim 5 wherein the expression of the isolated nucleic acid sequence allows the selected variant protein to be expressed from a host cell and bind to said target molecule.

### Claim 8 (depends on 5)

The isolated nucleic acid sequence of claim 5 is combined with a second nucleic acid sequence that encodes for a second protein having specific binding affinity to a second target molecule, and that expression of the combined nucleic acid sequences creates an oligomeric protein capable of binding to multi-target molecules at the same time.

### Claim 9 (depends on 1)

The protein of claim 1 further comprising at least one functional domain, linker sequence, sterically restricted intervening sequence, signal peptide, protease recognition site, and/or tag fused to the FHA domain.

### Claim 10 (depends on 9)

The protein of claim 9 wherein the functional domain is an additional FHA domain.

### Claim 11 (depends on 1)

The protein of claim 1 wherein the FHA domain is located within another functional domain.

### Claim 12 (depends on 1)

The protein of claim 1 wherein the specific binding affinity activates, inhibits, or enhances a biochemical pathway.

### Claim 13 (depends on 1)

The protein of claim 1, wherein the FHA domain is derived from SEQ ID No. 9.

### Claim 14

An artificial protein capable of recognizing an epitope wherein the artificial protein comprising:
a. An FHA domain derived from SEQ ID NO. 9 having a scaffold domain and loop domains; and
b. At least one mutated loop region or non-highly conserved sequence of the FHA domain,
c. The protein having specific binding affinity to a target molecule that does not include interactions with phosphate moieties based on naturally occurring, or in vivo FHA domain recognition sites.

### Claim 15 (depends on 14)

The protein of claim 14 further comprising at least one functional domain, linker sequence, sterically restricted intervening sequence, signal peptide, protease recognition site, and/or tag fused to the FHA domain.

### Claim 16 (depends on 15)

The protein of claim 15 wherein the functional domain is an additional FHA domain.

### Claim 17 (depends on 14)

The protein of claim 14 wherein the FHA domain is located within another functional domain.

### Claim 18 (depends on 14)

The protein of claim 14 wherein the specific binding affinity activates, inhibits, or enhances a biochemical pathway.

### Claim 19 (depends on 1)

The protein of claim 1 further comprising a multimer domain capable of forming multimeric proteins with at least one other protein molecule.

### Claim 20 (depends on 19)

The protein of claim 19 wherein the multimer domain comprises a CC1 domain.

### Claim 21 (depends on 14)

The protein of claim 14 further comprising a multimer domain capable of forming multimeric proteins with at least one other protein molecule.

### Claim 22 (depends on 21)

The protein of claim 21 wherein the multimer domain comprises a CC1 domain.