SRP and Sec pathway leader peptides for antibody phage display and antibody fragment production in E. coli.
Antibody phage display is a key technology for the generation of recombinant (human) antibodies for research, diagnostics and therapy. Most antibody fragments can only be folded correctly in the oxidizing environment of the periplasm of Escherichia coli. A multitude of leader peptides has been used for secretion of antibody::pIII fusion proteins into the periplasm, but a systematic study of their impact on the performance of antibody phage display systems has not been reported so far. In this work we have analysed the influence of various leader peptides on antibody phage display efficiency and production yields of soluble antibody fragments. Four leader peptides using the Sec pathway (PelB, OmpA, PhoA and pIII) and three using the SRP pathway (DsbA, TorT and TolB) were compared. Both pathways are compatible with antibody phage display and the production of soluble antibody fragments. The applicability of the SRP pathway to antibody phage display and the production of functional scFvs is shown here for the first time.
Thie
Holger
H
Technische Universität Braunschweig, Institut für Biochemie und Biotechnologie, Abteilung Biotechnologie, Spielmannstr. 7, 38106 Braunschweig, Germany.
Schirrmann
Thomas
T
Paschke
Matthias
M
Dübel
Stefan
S
Hust
Michael
M
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D000941:Antigens; D015153:Blotting, Western; D002645:Chickens; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D005822:Genetic Vectors; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis*; D008969:Molecular Sequence Data; D019151:Peptide Library; D021382:Protein Sorting Signals; D018271:Signal Recognition Particle / Q000378:metabolism*; D012995:Solubility
D000906:Antibodies; D000941:Antigens; D029968:Escherichia coli Proteins; D007128:Immunoglobulin Fragments; D019151:Peptide Library; D021382:Protein Sorting Signals; D018271:Signal Recognition Particle