Expression vector for secreting antibody fragment using
View Patent ↗A recombinant expression vector capable of expressing and secreting an antibody fragment fused with E. coli thermostable enterotoxin signal sequence derivative or E. coli outer membrane protein A (Omp A) signal sequence in the form of a soluble heterozygote protein is used to mass-produce the antibody fragment by culturing a microorganism transformed with the expression vector in a medium and collecting the antibody fragment secreted from the transformed microorganism into the medium.
1. A method for producing an antibody fragment, comprising the steps of:
1) preparing an expression vector comprising a gene encoding a light chain of the antibody fragment fused with E. coli thermostable enterotoxin signal sequence derivative and a gene encoding a heavy chain of the antibody fragment fused with E. coli outer membrane protein A signal sequence, wherein the expression of the genes encoding the light chain and the heavy chain is regulated by a single promoter;
2) transforming a microorganism with the expression vector;
3) culturing the transformed microorganism in a medium; and
4) collecting the antibody fragment from the medium or from the microorganism.
2. The method of claim 1 , wherein the antibody fragment is derived from a chimeric antibody, a humanized antibody or a human antibody.
3. The method of claim 1 , wherein the antibody fragment is selected from the group consisting of Fab, Fab′, F(ab′) 2 and scFv.
4. The method of claim 1 , wherein the E. coli thermostable enterotoxin signal sequence derivative has the nucleotide sequence of SEQ ID NO: 17 and the E. coli outer membrane protein A signal sequence has the nucleotide sequence of SEQ ID NO: 23.
5. The method of claim 1 , wherein the promoter is T7 promoter or Tac promoter.
6. The method of claim 1 , wherein the antibody fragment is a fragment of anti-tumor necrosis factor-alpha antibody.
7. The method of claim 1 , wherein the expression vector is pmsoDLHF_N/S.
8. The method of claim 1 , wherein the microorganism is E. coli.
9. The method of claim 8 , wherein the microorganism transformed with the expression vector is E. coli BL21/pmsoDLHF_N/S(HM10924) (KCCM-10513).
10. An expression vector comprising a gene encoding a light chain of the antibody fragment fused with E. coli thermostable enterotoxin signal sequence derivative and a gene encoding a heavy chain of the antibody fragment fused with E. coli outer membrane protein A signal sequence, wherein the expression of the genes encoding the light chain and the heavy chain is regulated by a single promoter, and the antibody fragment expressed from the expression vector is secreted into a culture medium where a host carrying the expression vector is cultured or into a periplasmic space of the host.
11. The expression vector of claim 10 , wherein the antibody fragment is derived from a chimeric antibody, a humanized antibody or a human antibody.
12. The expression vector of claim 10 , wherein the antibody fragment is selected from the group consisting of Fab, Fab′, F(ab′) 2 and scFv.
13. The expression vector of claim 10 , wherein the E. coli thermostable enterotoxin signal sequence derivative has the nucleotide sequence of SEQ ID NO: 17 and the E. coli outer membrane protein A signal sequence has the nucleotide sequence of SEQ ID NO: 23.
14. The expression vector of claim 10 , wherein the antibody fragment is a fragment of anti-tumor necrosis factor-alpha antibody.
15. The expression vector of claim 10 , wherein the promoter is T7 promoter or Tac promoter.
16. The expression vector of claim 15 , which is pmsoDLHF_N/S.
17. A microorganism transformed with the expression vector of claim 10 .
18. The microorganism of claim 17 , which is E. coli.
19. The microorganism of claim 18 , which is E. coli BL21/pmsoDLHF_N/S(HM10924) (KCCM-10513).