Process for the production of proteins and the production of arrays of proteins

• US 7,202,055 B2
• Filed: 12/19/2001
• Issued: 04/10/2007
• Est. Priority Date: 12/20/2000
• Status: Expired due to Fees

• Current Assignee: Altana Pharma AG (Takeda Pharmaceutical Company Limited)
• Original Assignee: Altana Pharma AG (Takeda Pharmaceutical Company Limited)
• Inventors: Schafer, Klaus Peter, Muhldorfer, Ingeborg
• Primary Examiner(s): Priebe; Scott D.
• Assistant Examiner(s): BURKHART, MICHAEL D
• Application Number:
  US10/399,954
• Publication Number:
  US 20040023329A1
• Time in Patent Office: 1,938 Days
• Field of Search: 435/69.1, 435/325, 435/252.33, 435/254.23
• US Class Current: 435/69.1
• CPC Class Codes: C12N 15/70 Vectors or expression systems specially adapted for E. coli

Abstract

Described is a novel process for the production of at least one protein of interest by secretion of the protein of interest from a pro- or eukaryotic host cell in a compartment system, which host cell is stably expressing a secretion system and capable of heterologous secretion of the protein of interest and which compartment system has at least a first and a second compartment and wherein the host cell is located in the first compartment and wherein the first and second compartment are separated from each other by a barrier, wherein the barrier is permeable for the secreted protein of interest, but not permeable for the host cell.

Claims

1. A process for the production of at least one protein of interest comprising culturing a host cell in a compartment system, which host cell is stably expressing a secretion system and capable of secreting the heterologous protein of interest, wherein the protein of interest is secreted by the host cell by means of a secretion signal, and which compartment system has at least a first and a second compartment, and wherein the host cell is located in the first compartment, and wherein the first and second compartment are separated from each other by a barrier, wherein the barrier is permeable for the secreted protein of interest, but not permeable for the host cell.

2. The process according to claim 1, wherein the host cell is expressing a secretion system encoded on a mobile vector.

3. The process according to claim 2, wherein the mobile vector is either a plasmid or a cosmid.

4. The process according to claim 1, wherein the barrier is part of the first compartment of the compartment system.

5. The process according to claim 1, wherein the barrier is a membrane filter, which membrane filter has a pore diameter that prevents host cell migration and allows the diffusion of the secreted protein of interest through the membrane.

6. The process according to claim 1, wherein the first compartment is located above the second compartment and the secreted protein of interest will diffuse by means of gravity through the barrier from the first compartment to the second compartment.

7. The process according to claim 5, wherein the host cell is located on the surface of the filter and the second compartment is a solid phase attached to the opposite surface of the filter and which solid phase is capable of retaining the secreted protein after diffusion through the filter.

8. The process according to claim 1, wherein the host cell is either E.coli or Pichia patoris.

9. The process according to claim 1, wherein the secretion of the protein of interest is effected by growth of the host cell.

10. The process according to claim 5, wherein the first and second compartment are stack microwells which are separated by said filter.

11. The process according to claim 1, wherein the protein of interest is obtained in the second compartment.

12. The process according to claim 1, wherein the secreted protein of interest carries an affinity tag, an epitope tag and/or carries a protease cleavage site suitable for removal of secretion signal and/or tag sequences.

13. The process according to claim 1 for the production of several different proteins of interest, wherein each host cell secreting a respective protein of interest is located in a defined area of the first compartment and each of the secreted proteins of interest will migrate to a defined area of the second compartment or wherein each host cell secreting a respective protein of interest is located in different first compartments and each of the secreted proteins of interest will migrate to different second compartments.

14. A process for the production of an array of several different proteins of interest, comprising culturing host cells in a compartment system, which host cells are stably expressing a secretion system, wherein the proteins of interest are secreted by the host cell by means of secretion signals, which enables the host cells to secrete the respective heterologous protein of interest and which compartment system has at least a first and a second compartment and wherein the different host cells are arranged in the form of an array in said first compartment and wherein the first and second compartment are separated from each other by a barrier, wherein the barrier is permeable for the secreted proteins of interest, but not permeable for the host cell and wherein the secreted proteins of interest are received in the second compartment in the form of an array.

15. The process according to claim 14, wherein the barrier is part of the first compartment of the compartment system.

16. The process according to claim 14, wherein the array of proteins corresponds to the proteome of an organism of interest.