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US 10,066,027 B2Granted
Protein production systems and methods thereof
AssigneeOhio State Innovation Foundation
InventorsDavid Webster Wood, Changhua Shi
Life of this patent
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Abstract
Disclosed herein is a protein purification system and methods of using the system. In particular, disclosed is a split intein comprising an N-terminal intein segment, which can be immobilized, and a C-terminal intein segment, which has the property of being self-cleaving, and which can be attached to a protein of interest. Through the self-cleaving mechanism of the intein, the protein of interest can be purified.
Claims19
A protein purification system, wherein the system comprises a split intein comprising two separate peptides:
- an N-terminal intein segment and a C-terminal intein segment, wherein the N-terminal intein segment comprises one or more amino acids at its C-terminus which allows for covalent immobilization of the N-terminal intein segment to a solid support, and wherein cleaving of the C-terminal intein segment is more sensitive to extrinsic conditions compared to a native intein, wherein a protein of interest (POI) is attached to the C-terminal intein segment at its C-terminus.
A method of purifying a protein of interest, the method comprising:
- a. utilizing a split intein comprising two separate peptides:
- an N-terminal intein segment and a C-terminal intein segment;
- b. covalently immobilizing the N-terminal intein segment to a solid support;
- c. attaching a protein of interest to the C-terminal intein segment, wherein cleaving of the C-terminal intein segment is more sensitive to extrinsic conditions compared to a native intein;
- d. exposing the N-terminal intein segment and the C-terminal intein segment to each other so that they associate;
- e. washing the solid support to remove non-bound material;
- f. placing the associated the N-terminal intein segment and the C-terminal intein segment under conditions that allow for the intein to self-cleave;
- g. isolating the protein of interest.
5 more elements
Dependencies are inferred from the claim text.