Method for enzymatic production of GLP-2(1-33) and GLP-2(1-34) peptides
AssigneeNPS Pharmaceuticals, Inc.
InventorsFred W. Wagner, Peng Luan, Yuannan Xia, Daniel Strydom, Jin Seog Seo
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Abstract
The invention provides methods for making peptides from a polypeptide containing at least one copy of the peptide using clostripain to excise the peptide from the polypeptide. The methods enable the use of a single, highly efficient enzymatic cleavage to produce any desired peptide sequence.
Claims19
An isolated nucleic acid encoding a polypeptide for production of a GLP-2 peptide, the encoded polypeptide comprising:
- two or more copies of the GLP-2 peptide;
- and
- a clostripain cleavage recognition sequence;
- wherein the clostripain cleavage recognition sequence is comprised of amino acids at the N-terminus and/or the C-terminus of the GLP-2 peptide and/or of an optional linker, the clostripain cleavage recognition sequence positioned to allow cleavage by clostripain between the copies of the GLP-2 peptide;
- wherein the clostripain cleavage recognition sequence comprises Formula I:
- -Xaa1-Xaa2-Xaa3- (I);
- wherein Xaa1 is aspartic acid, glycine, proline or glutamic acid, Xaa2 is arginine, and Xaa3 is not an acidic amino acid.
An isolated nucleic acid encoding a polypeptide for production of a GLP-2 peptide, the encoded polypeptide comprising:
- a GLP-2 peptide;
- wherein the GLP-2 peptide is selected from GLP-2(1-34) (SEQ ID NO:9), GLP-2(1-33) (SEQ ID NO:11), GLP-2(1-34,A2G) (SEQ ID NO:15) and GLP-2(1-33,A2G) (SEQ ID NO:13);
- and
- a clostripain cleavage recognition sequence;
- wherein the clostripain cleavage recognition sequence is comprised of amino acids at the N-terminus of the GLP-2 peptide and/or of an optional linker, the clostripain cleavage recognition sequence positioned to allow cleavage by clostripain at the N-terminus of the GLP-2 peptide;
- wherein the clostripain cleavage recognition sequence comprises Formula I:
- -Xaa1-Xaa2-Xaa3- (I);
- wherein Xaa1 is aspartic acid, glycine, proline or glutamic acid, Xaa2 is arginine, and Xaa3 is not an acidic amino acid.
An isolated nucleic acid encoding a polypeptide for production of a GLP-2 peptide, the encoded polypeptide comprising the following formula:
- Xaa1-Xaa2-Peptide
- wherein Xaa1 is aspartic acid, glycine, proline or glutamic acid, Xaa2 is arginine, and Peptide is a GLP-2 peptide;
- and
- wherein the GLP-2 peptide is selected from GLP-2(1-34) (SEQ ID NO:9, GLP-2(1-33) (SEQ ID NO:11), GLP-2(1-34,A2G) (SEQ ID NO:15) and GLP-2(1-33,A2G) (SEQ ID NO:13).
Dependencies are inferred from the claim text.