Recombinant NEL-like (NELL) protein production
View Patent ↗The present invention provides a method and system for producing a NELL protein. The method and system comprise a CELL encoding a NELL protein or peptide and a non-insect secretory signal peptide.
1. A method of expressing a peptide in a mammalian cell comprising:
providing a nucleic acid construct including at least a nucleic acid encoding at least a NEL-like protein (NELL) peptide in frame with a nucleic acid encoding a heterologous, non-insect secretory signal peptide;
transfecting a mammalian cell with the nucleic acid construct;
culturing the mammalian cell under conditions that permit expression of the NELL peptide;
collecting NELL peptide secreted from the cell; and
substantially purifying the NELL peptide using a chromatography process that uses membrane chromatography or affinity chromatography;
wherein the nucleic acid encodes NELL1 or NELL2, wherein NELL1 is selected from the group consisting of: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, and SEQ ID NO:6,
wherein NELL2 is selected from the group consisting of: SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, and SEQ ID NO:14.
2. The method of claim 1 , wherein the mammalian cell is a Chinese hamster ovary (CHO) cell.
3. The method of claim 1 , wherein substantially purifying comprises membrane chromatography.
4. The method of claim 3 , further comprising
testing the activity of the NELL peptide to induce bone formation.
5. The method of claim 1 , further comprising
testing the activity of the NELL peptide to induce bone formation.
6. The method of claim 1 , wherein substantially purifying comprises affinity chromatography.
7. The method of claim 1 , wherein the non-insect secretory signal peptide is a plant secretory signal peptide or an animal secretory signal peptide, wherein the animal secretory signal peptide is a mammalian secretory signal peptide.
8. A nucleic acid construct for expressing a NEL-like protein (NELL peptide) in a mammalian cell, the nucleic acid construct comprising at least a nucleic acid encoding at least a NELL peptide in frame with a nucleic acid encoding a heterologous non-insect signal peptide that is a non-insect secretory signal peptide;
wherein the nucleic acid encodes NELL1 or NELL2, wherein the NELL1 is selected from the group consisting of: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, and SEQ ID NO:6; and
wherein NELL2 is selected from the group consisting of: SEQ ID NO:7, SEQ ID NO: 8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, and SEQ ID NO:14.
9. The nucleic acid construct of claim 8 , wherein the mammalian cell is a CHO cell.
10. The nucleic acid construct of claim 8 , wherein the non-insect secretory signal peptide is a plant secretory signal peptide or an animal secretory signal peptide, wherein the animal secretory signal peptide is a mammalian secretory signal peptide.
11. A mammalian cell line for expressing a NEL-like protein (NELL) peptide, the cell line including a nucleic acid construct comprising at least a nucleic acid encoding at least a NELL peptide in frame with a nucleic acid encoding a heterologous non-insect signal peptide that is a non-insect secretory signal peptide;
wherein the nucleic acid encodes NELL or NELL2 peptide, wherein NELL1 is selected from the group consisting of: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, or SEQ ID NO:6; and
wherein NELL2 is selected from the group consisting of: SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, and SEQ ID NO:14.
12. The cell of claim 11 , wherein the mammalian cell line comprises a CHO cell.
13. The cell of claim 12 , wherein the CHO cell secretes NELL1 or NELL2 peptide.
14. The cell line of claim 11 , wherein the non-insect secretory signal peptide is a plant secretory signal peptide or an animal secretory signal peptide, and wherein the animal secretory signal peptide is a mammalian secretory signal peptide.