Vectors, methods, systems and kits for protein purification
AssigneeNational Chiao Tung University
InventorsYaw-Kuen Li, Yue-Jin Wu
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Abstract
Disclosed herein are autocleaved peptide linkers for producing purified proteins. The autocleaved peptide linkers are inserted between a chitin binding protein (CBP) and a target protein to form a fusion protein. Upon expression of the fusion protein, it is allowed to pass a chitin matrix so that the CBP portion of the fusion protein may be bound with the chitin matrix, the peptide linker then undergoes auto-cleavage in a buffer solution at a pH value of about 5.5-7.5 to release the target protein. The chitin matrix may be regenerated with another buffer solution at a pH value of about 3-4, that is, to release the bound CBP and return to its unbound form. The chitin matrix may be reused for at least 6 times without losing its function.
Claims18
A vector for producing a fusion protein, encoded by a polynucleotide sequence, comprising:
- a promoter;
- a first polynucleotide operably linked to the promoter and encodes a chitin-binding-protein (CBP);
- a second polynucleotide operably linked to the first polynucleotide and encodes a linker, which upon expression produces a peptide having an amino acid sequence selected from the group consisting of SEQ ID NO:
- 1, SEQ ID NO:
- 2, SEQ ID NO:
- 3 and SEQ ID NO:
- 4, and the peptide being auto-cleavaging under pH value between about 5.5 to about 7.5 without subjecting to a protease treatment;
- and
- a third polynucleotide operably linked to the second polynucleotide and encodes a target protein.
Dependencies are inferred from the claim text.