A single-chain antibody fragment is functionally expressed in the cytoplasm of both Escherichia coli and transgenic plants

TitleA single-chain antibody fragment is functionally expressed in the cytoplasm of both Escherichia coli and transgenic plants
Publication TypeArticolo su Rivista peer-reviewed
Year of Publication1999
AuthorsTavladoraki, P., Girotti A., Donini Marcello, Arias F.J., Mancini C., Morea V., Chiaraluce R., Consalvi V., and Benvenuto E.
JournalEuropean Journal of Biochemistry
Volume262
Pagination617-624
ISSN00142956
KeywordsAntibody, article, Cysteine, cytoplasm, Disulfides, Escherichia coli, Genetically Modified, immunoglobulin variable region, nonhuman, oxidation reduction state, Plants, priority journal, protein analysis, Protein Denaturation, protein expression, Protein Folding, Protein Sorting Signals, protein stability, transgenic plant
Abstract

Despite the well-known crucial role of intradomain disulfide bridges for immunoglobulin folding and stability, the single-chain variable fragment of the anti-viral antibody F8 is functionally expressed when targeted to the reducing environment of the plant cytoplasm. We show here that this antibody fragment is also functionally expressed in the cytoplasm of Escherichia coli. A gel shift assay revealed that the single-chain variable fragment (scFv) accumulating in the plant and bacterial cytoplasm bears free sulfhydryl groups. Guanidinium chloride denaturation/renaturation studies indicated that refolding occurs even in a reducing environment, producing a functional molecule with the same spectral properties of the native scFv(F8). Taken together, these results suggest that folding and functionality of this antibody fragment are not prevented in a reducing environment. This antibody fragment could therefore represent a suitable framework for engineering recombinant antibodies to be targeted to the cytoplasm.

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URLhttps://www.scopus.com/inward/record.uri?eid=2-s2.0-0033152614&doi=10.1046%2fj.1432-1327.1999.00443.x&partnerID=40&md5=d4118b32bba39f18e54d6a9b9f2b81bb
DOI10.1046/j.1432-1327.1999.00443.x