Capture purification processes for proteins expressed in a non-mammalian system
Methods of purifying proteins expressed in non-mammalian expression systems in a non-native soluble form directly from cell lysate are disclosed. Methods of purifying proteins expressed in non-mammalian expression systems in a non-native limited solubility form directly from a refold solution are also disclosed. Resin regeneration methods are also provided.
1. A method of purifying a protein expressed in a non-native soluble form in a non-mammalian expression system, the method comprising:
(a) lysing a non-mammalian cell in which the protein is expressed in a nonnative soluble form to generate a cell lysate;
(b) filtering the cell lysate;
(c) contacting the filtered cell lysate with a separation matrix under conditions suitable for the protein to associate with the separation matrix;
(d) washing the separation matrix;
(e) eluting the protein from the separation matrix; and
(f) isolating the protein after elution.
2. The method of claim 1 , wherein the non-mammalian expression system comprises bacteria cells.
3. The method of claim 2 , further comprising refolding the isolated protein to its native form.
4. The method of claim 2 , wherein the separation matrix is a nonaffinity resin selected from the group consisting of: ion exchange, mixed mode, and a hydrophobic interaction resin.
5. The method of claim 2 , wherein the separation matrix is an affinity resin selected from the group consisting of: Protein A, Protein G, and a synthetic mimetic affinity resin.
6. The method of claim 2 , wherein the protein is a complex protein.
7. The method of claim 6 , wherein the complex protein is selected from the group consisting of: a multimeric protein, an antibody, and an Fc fusion protein.
8. The method of claim 1 , wherein the non-mammalian expression system comprises yeast cells.
9. The method of claim 8 , further comprising refolding the isolated protein to its native form.
10. The method of claim 8 , wherein the separation matrix is a nonaffinity resin selected from the group consisting of: ion exchange, mixed mode, and a hydrophobic interaction resin.
11. The method of claim 8 , wherein the separation matrix is an affinity resin selected from the group consisting of: Protein A, Protein G, and a synthetic mimetic affinity resin.
12. The method of claim 8 , wherein the protein is a complex protein.
13. The method of claim 12 , wherein the complex protein is selected from the group consisting of: a multimeric protein, an antibody, and an Fc fusion protein.
14. The method of claim 1 , further comprising refolding the isolated protein to its native form.
15. The method of claim 14 , wherein the protein is a complex protein.
16. The method of claim 15 , wherein the complex protein is selected from the group consisting of: a multimeric protein, an antibody, and an Fc fusion protein.
17. The method of claim 14 , wherein the separation matrix is a nonaffinity resin selected from the group consisting of: ion exchange, mixed mode, and a hydrophobic interaction resin.
18. The method of claim 14 , wherein the separation matrix is an affinity resin selected from the group consisting of: Protein A, Protein G, and a synthetic mimetic affinity resin.