Eukaryotic signal sequences for polypeptide expression and polypeptide display libraries
View Patent ↗The present invention generally relates to methods and compositions for expressing proteins or polypeptides in prokaryotic hosts using eukaryotic signal sequences.
1. A method of preparing a polypeptide display library, which method comprises the steps of introducing a polynucleotide encoding an exogenous polypeptide into the genome of a replicable genetic package to form a fusion protein with an endogenous protein that is normally expressed from the outersurface of the replicable genetic package, wherein the polynucleotide is operably linked to a eukaryotic signal sequence which encodes a signal peptide MGALAVFAVACLAAVASVAHA (SEQ ID NO: 3), and expressing said fusion protein, wherein the replicable genetic package is a bacteriophage or phagemid.
2. A method according to claim 1 , wherein the polypeptide is a eukaryotic polypeptide.
3. A method according to claim 1 , wherein the polypeptide is a prokaryotic polypeptide.
4. A method according to claim 1 , wherein the polypeptide is an antibody.
5. A method according to claim 1 , wherein the polypeptide is an antibody that is a mouse antibody, a human antibody, or a chimeric antibody.
6. A method according to claim 1 , wherein the polypeptide is a whole antibody or binding fragment thereof.
7. A method according to claim 1 , wherein the polypeptide is an antibody binding fragment that is a single chain fragment, an Fv fragment or a Fab fragment.
8. A method according to claim 1 , wherein the signal sequence is 5′ to the polynucleotide sequence encoding the polypeptide.
9. A method according to claim 1 , wherein the signal sequence and the polypeptide are separated by zero amino acids.
10. A method according to claim 1 , wherein the bacteriophage is a filamentous phage.
11. A method according to claim 10 , wherein the filamentous phage is an M13, Fd, or F1 phage.
12. A method according to claim 1 , further comprising the steps of (a) screening for the polypeptide, (b) enriching the polypeptide display library, and/or (c) subcloning the polynucleotide encoding the identified polypeptide into an expression vector.