Negative effects of a disulfide bond mismatch in anti-rabies G protein single-chain antibody variable fragment FV57

作者：Duan, Ye; Gu, Tiejun; Zhang, Xizhen; Jiang, Chunlai; Yuan, Ruosen; Li, Zhuang; Wang, Dandan; Chen, Xiaoxu; Wu, Chunlai; Chen, Yan; Wu, Yongge; Kong, Wei^*

来源：Molecular Immunology, 2014, 59(2): 136-141.

DOI：10.1016/j.molimm.2014.01.006

• Rabies virus
• Glycoprotein
• scFv
• Disulfide bond mismatch

Rabies virus (RV) causes a fatal infectious disease requiring efficient post-exposure prophylaxis (PEP), which includes a rabies vaccine and rabies immunoglobulin (RIG). The single-chain antibody variable fragment (scFv), a small engineered antibody fragment derived from an antibody variable heavy chain and light chain, has the potential to replace the current application of RIG. In previous studies, we constructed and evaluated an anti-rabies virus G protein scFv (FV57) based on the monoclonal antibody CR57. Of the five cysteines in FV57, four are linked in intra-chain disulfide bonds (Cys-V(H)28/Cys-V(H)98 and Cys-V(L)16/Cys-V(L)84), and one is free (Cys-V(L)85). However, the thiol in Cys-V(L)85 neighboring Cys-V(L)84 in the CDR3 of the light chain is likely to mismatch with the thiol in Cys-V(L)16 during the renaturing process. In order to study effects of the mismatched disulfide bond, Cys-V(L)85 and Cys-V(L)84 of FV57 were mutated to serine to construct mutants FV57(VL85S) and FV57(VL84S). Furthermore, the disulfide bonds in the light chain of FV57, FV57(VL85S) and FV57(VL84S) were deleted by mutating Cys-V(L)16 to serine. All mutants were prepared and evaluated along with the original FV57. The results indicated that the mismatched disulfide bond of FV57 linking the light chain FR1 and CDR3 would confer deleterious negative effects on its activity against RV, likely due to spatial hindrance in the light chain CDR3. Moreover, avoidance of the disulfide bond mismatch provided an additional 30% protective efficacy against RV infection in the mouse RV challenge model. Thus, modifications of FV57 to eliminate the disulfide bond mismatch may provide a candidate therapeutic agent for effective PEP against rabies.

• 出版日期2014-6
• 单位吉林大学