Increased dynamic binding capacity in ion exchange chromatography by addition of polyethylene glycol
View Patent ↗The present invention generally relates to novel processes for protein purification in high salt solutions such as cell culture broth by increasing the dynamic binding capacity of a resin with the addition of polyethylene glycol.
1. A process for protein purification by ion exchange chromatography from a protein solution having a salt concentration sufficient to significantly reduce the binding of the protein to the ion exchange resin, the process, comprising the steps of:
a. adding from about 0.5% w/v to about 6% w/v polyethylene glycol (PEG) to a protein solution having a salt concentration sufficient to significantly reduce the binding of the protein to an ion exchange resin to form a protein-PEG solution; and,
b. binding the protein to the ion exchange resin by contacting the ion exchange resin with the protein-PEG solution.
2. The process of claim 1 , wherein the protein solution is a cell culture broth.
3. The process of claim 1 , wherein the PEG is chosen with a molecular weight between about PEG 400 and about PEG 10000, and mixtures thereof.
4. The process of claim 1 , wherein the PEG is PEG 4600.
5. The process of claim 1 , wherein the ion exchange resin is a cationic exchange resin.
6. The process of claim 5 , wherein the conductivity of the protein-PEG solution is greater than about 9 mS/cm.
7. The process of claim 1 , wherein the ion exchange resin is an anionic exchange resin.
8. The process of claim 7 , wherein the conductivity of the protein-PEG solution is greater than about 11 mS/cm.
9. The process of claim 1 , wherein the binding of the protein to the ion exchange resin occurs about a column.
10. The process of claim 1 , wherein the step of contacting is at a pH in a range of about 4.0 to about 10.0.
11. The process of claim 1 , wherein the protein solution is diluted.
12. The process of claim 5 , wherein the ion exchange resin is a cation exchanger at a pH of about 5.0.
13. The process of claim 1 , wherein the protein is selected from the group consisting of bovine γ-globulin, bovine serum albumin, and lysozyme.
14. The process of claim 1 , further comprising determining an ideal concentration of PEG for a particular protein by adding progressively greater concentrations of PEG to facilitate binding during loading of the resin until an acceptable amount of precipitation is observed.
15. The process of claim 1 , further comprising filtering the protein-PEG solution.
16. The process of claim 1 , wherein the step of binding the protein to the ion exchange resin is preceded by a step of equilibrating the ion exchange resin with a binding buffer.
17. A process for the separation of a protein of interest by ion exchange chromatography from a cell culture broth having a salt concentration sufficient to significantly reduce the binding of the protein to the ion exchange resin, the process comprising the steps of:
a. adding polyethylene glycol (PEG) to the cell culture broth to form a PEG-cell culture broth;
b. binding the protein of interest to the ion exchange resin by contacting the ion exchange resin with the PEG-cell culture broth; and,
c. separating the protein of interest from the PEG-cell culture broth.
18. The process of claim 17 , wherein the protein of interest is a recombinant protein.
19. The process of claim 17 , wherein the ion exchange resin is either a cation exchange resin or an anion exchange resin.
20. The process of claim 17 , further comprising the step of determining an ideal concentration of PEG for a particular protein by adding progressively greater concentrations of PEG to facilitate binding during loading of the resin until an acceptable amount of precipitation is noted.
21. A mixture comprising an ion exchange resin and a solution comprising a cell culture broth, polyethylene glycol (PEG) in an amount which is between about 1% w/v and 8% w/v of the cell culture broth, and a protein;
wherein
the protein is bound to the ion exchange resin, and
the cell culture broth has a salt concentration sufficient to significantly reduce the binding of the protein to the ion exchange resin in the absence of the PEG.
22. The mixture of claim 21 , wherein the ion exchange resin is a cationic ion exchange resin and the conductivity of the solution is greater than about 9.
23. The mixture of claim 21 , wherein the ion exchange resin is an anionic ion exchange resin and the conductivity of the solution is greater than about 11.