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

Abstract

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