Method for cleavage of fusion proteins
AssigneeSemBioSys Genetics Inc.
InventorsGijs Van Rooijen, Joenel Alcantara, Maurice M. Moloney
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Abstract
An improved method for recovering recombinantly produced polypeptides is described. The method involves expressing the recombinant polypeptide as a fusion protein with a pro-peptide. The pro-peptide-polypeptide fusion protein can be cleaved and the recombinant polypeptide released under the appropriate conditions.
Claims53
A method for the preparation of a recombinant polypeptide comprising
- a) introducing into a host cell an expression vector comprising:
- (1) a nucleic acid sequence capable of regulating transcription in a host cell, operatively linked to
- (2) a chimeric nucleic acid sequence encoding a fusion protein, the chimeric nucleic acid sequence comprising (a) a nucleic acid sequence encoding a pro-peptide derived from an autocatalytically maturing zymogen, linked in reading frame to (b) a nucleic acid sequence heterologous to the pro-peptide and encoding the recombinant polypeptide;
- operatively linked to
- (3) a nucleic acid sequence encoding a termination region functional in said host cell,
- b) growing the host cell to produce said fusion protein;
- and
- c) altering the environment of the fusion protein so that the pro-peptide is cleaved from the fusion protein to release the recombinant polypeptide.
A chimeric nucleic acid sequence encoding a fusion protein comprising (a) a nucleic acid sequence encoding a pro-peptide from an autocatalytically maturing zymogen and (b) a nucleic acid sequence encoding a polypeptide that is heterologous to the pro-peptide.
A method of delivering a therapeutic or nutritional polypeptide to a human or animal comprising
- (a) providing a fusion protein comprising
- (i) a pro-peptide derived from an autocatalytically maturing enzyme, linked to
- (ii) a polypeptide that is heterologous to the pro-peptide and is a therapeutic or nutritional protein;
- and
- (b) administering the fusion protein to the human or animal where the therapeutic or nutritional polypeptide is cleaved from the pro-peptide.
A pharmaceutical composition comprising a fusion protein which comprises (a) a pro-peptide derived from an autocatalytically maturing zymogen and (b) a polypeptide that is heterologous to the pro-peptide, in admixture with a suitable diluent or carrier.
A food composition comprising a fusion protein which comprises a pro-peptide derived from an autocatalytically maturing zymogen and (b) a polypeptide that is heterologous to the pro-peptide, in admixture with a suitable diluent or carrier.
A pharmaceutical composition comprising a chimeric nucleic acid sequence encoding a fusion protein, the chimeric nucleic acid sequence comprising (a) a first nucleic acid sequence encoding a pro-peptide derived from an autocatalytically maturing zymogen and (b) a second nucleic acid sequence encoding a polypeptide that is heterologous to the pro-peptide.
A food composition comprising a chimeric nucleic acid sequence encoding a fusion protein, the chimeric nucleic acid sequence comprising (a) a first nucleic acid sequence encoding a pro-peptide derived from an autocatalytically maturing zymogen and (b) a second nucleic acid sequence encoding a polypeptide that is heterologous to the pro-peptide.
A fusion protein comprising (a) a pro-peptide derived from an autocatalytically maturing zymogen and (b) a polypeptide that is heterologous to the pro-peptide.
A use of a fusion protein comprising (i) a pro-peptide derived from an autocatalytically maturing enzyme, linked to (ii) a polypeptide that is heterologous to the pro-peptide and is a therapeutic or nutritional protein;
- to deliver a therapeutic or nutritional protein to a human or animal.
A polypeptide having pro-peptide activity and comprising the amino acid sequence of KLIP 11 (SEQ. ID. NO.:
- 7).
A polypeptide having pro-peptide activity and comprising the amino acid sequence of KLIP 12 (SEQ.ID. NO.:8).
A polypeptide having pro-peptide activity and comprising the amino acid sequence of KLIP 14 (SEQ.ID. NO.:9).
A polypeptide having pro-peptide activity and comprising the amino acid sequence of KLIP 15 (SEQ.ID. NO.:10).
Dependencies are inferred from the claim text.