Fusion proteins and methods of producing same
The present invention provides compositions and methods for treating or preventing hyperacute rejection.
1. A method of producing a mucin-immunoglobulin fusion polypeptide comprising:
a) providing a cell comprising:
i) a nucleic acid encoding a mucin polypeptide linked to a nucleic acid encoding at least a region of an immunoglobulin polypeptide;
ii) a nucleic acid encoding an α1,3 galactosyltransferase polypeptide; and
iii) a nucleic acid encoding a β 1,6,-N-acetylglucosaminyltransferase polypeptide;
b) culturing the cell under conditions that permit production of said mucin-immunoglobulin fusion polypeptide wherein said fusion polypeptide is glycosylated by said α1,3 galactosyltransferase and said β 1,6,-N-acetylglucosaminyltransferase; and
c) isolating said mucin-immunoglobulin fusion polypeptide.
2. The method of claim 1 , wherein the mucin polypeptide is selected from the group consisting of PSGL-1, MUC1, MUC2, MUC3, MUC4, MUC5a, MUC5b, MUC5c, MUC6, MUC11, MUC12, CD34, CD43, CD45, CD96, GlyCAM-1, and MAdCAM 3 or fragment thereof.
3. The method of claim 1 , wherein the mucin polypeptide comprises at least a region of a P-selectin glycoprotein ligand- 1.
4. The method of claim 1 , wherein said mucin polypeptide comprises an extracellular region of a P-selectin glycoprotein ligand- 1.
5. The method of claim 1 , wherein the immunoglobulin polypeptide comprises a region of a heavy chain immunoglobulin polypeptide.
6. The method of claim 1 , wherein the immunoglobulin polypeptide comprises an Fc region of an immunoglobulin heavy chain.
7. The method of claim 1 , wherein the cell is a eukaryotic cell, or a prokaryotic cell.
8. The method of claim 7 , wherein the eukaryotic cell is a mammalian cell, an insect cell or a yeast cell.
9. The method of claim 7 , wherein the prokaryotic cell is a bacterial cell.
10. The method of claim 7 , wherein the eukaryotic cell is a CHO cell, a COS cell or a 293 cell.
11. A method of producing a mucin-immunoglobulin fusion polypeptide comprising:
a) introducing into a cell
i) a nucleic acid encoding a mucin polypeptide linked to a nucleic acid encoding at least a region of an immunoglobulin polypeptide,
ii) a nucleic acid encoding an α1,3 galactosyltransferase polypeptide; and
iii) a nucleic acid encoding a β 1,6,-N-acetylglucosaminyltransferase polypeptide;
b) culturing the cell under conditions that permit production of said mucin-immunoglobulin fusion polypeptide wherein said fusion polypeptide is glycosylated by said α1,3 galactosyltransferase polypeptide and said β 1,6,-N-acetylglucosaminyltransferase; and
c) isolating said mucin-immunoglobulin fusion polypeptide.
12. A cell comprising:
a) a nucleic acid encoding a mucin polypeptide linked to a nucleic acid encoding at least a region of an immunoglobulin polypeptide,
b) a nucleic acid encoding an α1,3 galactosyltransferase polypeptide;
c) and a nucleic acid encoding a β 1,6,-N-acetylglucosaminyltransferase.
13. The cell of claim 12 , wherein the cell is a CHO cell.