Enhanced intracellular stability of sFv-Fc fusion intrabodies.
The ability of intracellular antibodies (intrabodies) to block the function of a target protein can be dependent on the stability of the single-chain antibody (sFv) when expressed in the intracellular environment. Low-affinity sFvs capable of reaching high steady-state levels can be more effective modulators of target proteins than high-affinity, unstable sFvs. In an effort to enhance the intracellular stability of sFvs when expressed as intrabodies, we have generated novel sFv-Fc fusion intrabodies. Fusion of the native sFv sequence with the entire heavy chain constant region fragment of IgG results in decreased turnover of the intrabody and enhanced steady-state accumulation of sFv-Fc protein, while maintaining the ability to target intrabody expression to sub-cellular compartments. Here, we describe the rationale and design for this strategy using two anti-cyclin E sFvs constructed for use as intrabodies.
Department of Cancer Biology, Wake Forest University Graduate School of Arts and Sciences, Winston-Salem, NC 27157, USA.
Randall W
RW
Strube
Si-Yi
SY
Chen
D045744:Cell Line, Tumor; D003001:Cloning, Molecular / Q000379:methods; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000235:genetics* / Q000378:metabolism*; D007135:Immunoglobulin Variable Region / Q000235:genetics* / Q000378:metabolism*; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism*
D007141:Immunoglobulin Fc Fragments; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins