التفاصيل البيبلوغرافية
العنوان: |
METHOD FOR UNIVERSAL ENZYMATIC PRODUCTION OF BIOACTIVE PEPTIDES |
Document Number: |
20120156719 |
تاريخ النشر: |
June 21, 2012 |
Appl. No: |
12/354053 |
Application Filed: |
January 15, 2009 |
مستخلص: |
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. |
Inventors: |
Wagner, Fred W. (Walton, NE, US); Luan, Peng (Omaha, NE, US); Xia, Yuannan (Lincoln, NE, US); Strydom, Daniel (Lincoln, NE, US); Merrifield, Edwin H. (Lynnwood, WA, US); Bossard, Mary J. (Madison, AL, US); Holmquist, Barton (Eagle, NE, US); Seo, Jin Seog (Brampton, CA) |
Claim: |
1-2. (canceled) |
Claim: |
3. A method for producing a desired peptide having a C-terminal acidic, aliphatic or aromatic amino acid from a polypeptide, comprising: combining clostripain and the polypeptide comprising formula (III): (Linker-Xaa3-Peptide1)n-Linker-Xaa3-Peptide1 (III); wherein the desired peptide comprises Xaa3-Peptide1; Peptide1 is any amino acid sequence other than Xaa1-Xaa2; n is an integer ranging from 0 to 50; Xaa3 is not an acidic amino acid; Linker is a cleavable peptide linker having Formula (IV): (Peptide5)m—Xaa1-Xaa2 (IV); wherein m is an integer ranging from 0 to 50; Xaa1 is aspartic acid, glycine, proline or glutamic acid; Xaa2 is arginine; and Peptide5 is any single amino acid residue or amino acid sequence not containing Xaa1-Xaa2. |
Claim: |
4. The method of claim 1, wherein the polypeptide is a soluble polypeptide. |
Claim: |
5. The method of claim 1, wherein the cleavage is performed at about 18° C. to about 25° C. |
Claim: |
6. The method of claim 1, wherein the cleavage is performed between a pH of about 5 to about 11. |
Claim: |
7. The method of claim 1, wherein the concentration of clostripain is about 0.01 to about 3.0 units of clostripain per about 2 to about 5 mg polypeptide. |
Claim: |
8. The method of claim 1, wherein the cleavage is performed in the presence of about 0.5 mM to about 10 mM CaCl2. |
Claim: |
9. The method of claim 3, wherein the Linker comprises Pro-Gly-Xaa1-Xaa2; and Xaa1 is aspartic acid. |
Claim: |
10. The method of claim 3, wherein the Linker comprises Val-Asp-Xaa1-Xaa2; and Xaa1 is aspartic. |
Claim: |
11. The method of claim 3, wherein the Linker comprises Ile-Thr-Xaa1-Xaa2; and Xaa1 is aspartic acid. |
Claim: |
12. The method of claim 3, wherein Peptide5 is selected from the group consisting of Gly-Ser-, Cys-His-, Cys-His-Xaa-Xaa- and Val-Asp-; Xaa1 is Aspartic acid or Glutamic acid; Xaa independently is any amino acid residue other than Arginine and m is an integer of 1 to 10. |
Claim: |
13. The method of claim 1, wherein the desired peptide comprises anyone of SEQ ID NO:1-16. |
Claim: |
14. The method of claim 1, wherein the polypeptide comprises a soluble four copy or six copy GLP-2(1-34) sequence. |
Claim: |
15-71. (canceled) |
Current U.S. Class: |
435/681 |
Current International Class: |
12 |
رقم الانضمام: |
edspap.20120156719 |
قاعدة البيانات: |
USPTO Patent Applications |