title: A novel disulfide-stabilized single-chain variable antibody fragment against rabies virus G protein with enhanced in vivo neutralizing potency.
abstract: Rabies is a fatal infectious disease requiring efficient protection provided by post-exposure prophylaxis (PEP) with rabies immunoglobulin (RIG). The single-chain Fv fragment (scFv) is a small engineered antigen binding protein derived from antibody variable heavy (V(H)) and light (V(L)) chains. This novel antibody format may potentially replace the current application of RIG to detect and neutralize rabies virus (RV). However, the broad use of scFvs is confined by their generally low stability. In this study, a scFv (FV57) was constructed based on the monoclonal antibody, MAB57, against RV. To enhance its stability and neutralizing potency, a disulfide-stabilized scFv, ds-FV57, was also derived by introduction of cysteines at V(H)44 and V(L)100. Furthermore, the cysteine at V(L)85 of ds-FV57 was mutated to serine to construct ds-FV57(VL85Ser) in order to avoid potential mis-formed disulfide bonds which would alter the affinity of the scFv. The stability and activity of all three proteins expressed in Escherichia coli were evaluated. All of the constructed scFvs could provide efficient protection against RV infection both in vivo and in vitro. However, the stability of ds-FV57(VL85Ser) was notably improved, and its in vitro neutralizing potency against RV infection was enhanced. Our findings from these stabilization modifications support the feasibility of developing scFvs for PEP treatment of rabies.
authors.affiliation: National Engineering Laboratory for AIDS Vaccine, College of Life Science, Jilin University, Changchun 130012, China.
authors.forename: Ye
authors.initials: Y
authors.lastname: Duan
authors.forename: Tie-jun
authors.initials: TJ
authors.lastname: Gu
authors.forename: Chun-lai
authors.initials: CL
authors.lastname: Jiang
authors.forename: Ruo-sen
authors.initials: RS
authors.lastname: Yuan
authors.forename: Hua-fei
authors.initials: HF
authors.lastname: Zhang
authors.forename: Hong-jia
authors.initials: HJ
authors.lastname: Hou
authors.forename: Xiang-hui
authors.initials: XH
authors.lastname: Yu
authors.forename: Yan
authors.initials: Y
authors.lastname: Chen
authors.forename: Yong
authors.initials: Y
authors.lastname: Zhang
authors.forename: Yong-ge
authors.initials: YG
authors.lastname: Wu
authors.forename: Wei
authors.initials: W
authors.lastname: Kong
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology; D000915:Antibody Affinity; D006224:Cricetinae; D051379:Mice; D056990:Post-Exposure Prophylaxis / Q000379:methods*; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods; D055550:Protein Stability; D011818:Rabies / Q000517:prevention & control*; D011819:Rabies Vaccines / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*; D014759:Viral Envelope Proteins / Q000276:immunology*
chemical_list: D057134:Antibodies, Neutralizing; D011819:Rabies Vaccines; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; D014759:Viral Envelope Proteins