title: Effects of signal sequence on phage-displayed disulfide-stabilized single chain antibody variable fragment (sc-dsFv) libraries.
abstract: Phage-displayed single chain variable fragment (scFv) libraries are powerful tools in antibody engineering. Disulfide-stabilized scFv (sc-dsFv) with an interface disulfide bond is structure-wise more stable than the corresponding scFv. A set of recently discovered signal sequences replacing the wild type (pelB) signal peptidase cleavage site in the c-region has been shown to be effective in rescuing the expression of sc-dsFv libraries on the phage surface. However, the effects of the other regions of the signal sequence on the expression of the sc-dsFv libraries and on the formation of the interface disulfide bond in the phage-displayed sc-dsFv have not been clear. In this work, selected novel signal sequence variants in the h-region were shown to be equally effective in promoting sc-dsFv library expression on the phage surface; the expression level and complexity of the sc-dsFv libraries were comparable to the corresponding scFv libraries produced with the wild-type (pelB) signal sequence. The interface disulfide bond in the phage-displayed sc-dsFv was proven to form to a large extent in the library variant ensemble generated with signal sequence variants in both the h-region and the c-region. The sc-dsFv engineering platform established in this work can be applied to many of the known scFv molecules which are in need of a more stable version for the applications under harsh conditions or for longer shelf-life.
authors: Genomics Research Center, Academia Sinica, Taipei 115, Taiwan, ROC.
authors: Yu-Ching
authors: YC
authors: Lee
authors: Ing-Chien
authors: IC
authors: Chen
authors: Chung-Ming
authors: CM
authors: Yu
authors: Yi-Jen
authors: YJ
authors: Huang
authors: Hung-Ju
authors: HJ
authors: Hsu
authors: An-Suei
authors: AS
authors: Yang
mesh_terms: D000595:Amino Acid Sequence; D003545:Cysteine / Q000737:chemistry*; D006801:Humans; D008969:Molecular Sequence Data; D019151:Peptide Library; D015202:Protein Engineering; D021382:Protein Sorting Signals; D055550:Protein Stability; D057127:Single-Chain Antibodies / Q000737:chemistry* / Q000235:genetics* / Q000276:immunology; D042461:Vascular Endothelial Growth Factor A / Q000037:antagonists & inhibitors / Q000276:immunology*
chemical_list: D019151:Peptide Library; D021382:Protein Sorting Signals; D057127:Single-Chain Antibodies; D042461:Vascular Endothelial Growth Factor A; D003545:Cysteine