Fusion Peptide Improves Stability and Bioactivity of Single Chain Antibody against Rabies Virus = Fusion Peptide Improves Stability and Bioactivity of Single Chain Antibody against Rabies Virus

다국어 초록 (Multilingual Abstract)

The combination of rabies immunoglobulin (RIG) with a vaccine is currently effective against rabies infections, but improvements are needed. Genetic engineering antibody technology is an attractive approach for developing novel antibodies to replace RIG. In our previous study, a single-chain variable fragment, scFv57R, against rabies virus glycoprotein was constructed. However, its inherent weak stability and short half-life compared with the parent RIG may limit its diagnostic and therapeutic application. Therefore, an acidic tail of synuclein (ATS) derived from the C-terminal acidic tail of human alpha-synuclein protein was fused to the Cterminus of scFv57R in order to help it resist adverse stress and improve the stability and halflife. The tail showed no apparent effect on the preparation procedure and affinity of the protein, nor did it change the neutralizing potency in vitro. In the ELISA test of molecular stability, the ATS fusion form of the protein, scFv57R-ATS, showed an increase in thermal stability and longer half-life in serum than scFv57R. The protection against fatal rabies virus challenge improved after fusing the tail to the scFv, which may be attributed to the improved stability. Thus, the ATS fusion approach presented here is easily implemented and can be used as a new strategy to improve the stability and half-life of engineered antibody proteins for practical applications.

참고문헌 (Reference)

1 Koprowski H, "Use of hyperimmune antirabies serum concentrates in experimental rabies" 8 : 412-420, 1950

2 Lee EN, "Stabilizing peptide fusion for solving the stability and solubility problems of therapeutic proteins" 22 : 1735-1746, 2005

3 Bird RE, "Single-chain antigen-binding proteins" 242 : 423-426, 1988

4 Knobel DL, "Re-evaluating the burden of rabies in Africa and Asia" 83 : 360-368, 2005

5 Bourhy H, "Rabies, still neglected after 125 years of vaccination" 4 : e839-e839, 2010

6 Jiang H, "Purification of clinical-grade disulfide stabilized antibody fragment variable-Pseudomonas exotoxin conjugate (dsFv-PE38) expressed in Escherichia coli" 97 : 621-632, 2013

7 Yuan R, "Preparation and diagnostic use of a novel recombinant single-chain antibody against rabies virus glycoprotein" 98 : 1547-1555, 2014

8 Both L, "Passive immunity in the prevention of rabies" 12 : 397-407, 2012

9 Cai K, "Novel human 3-domain disulfide-stabilized antibody fragment against glycoprotein of rabies virus" 10 : 548-555, 2008

10 Goldman ER, "Negative tail fusions can improve ruggedness of single domain antibodies" 95 : 226-232, 2014

11 Duan Y, "Negative effects of a disulfide bond mismatch in anti-rabies G protein single-chain antibody variable fragment FV57" 59 : 136-141, 2014

12 Nagarajan T, "Monoclonal antibodies for the prevention of rabies: theory and clinical practice" 4 : 1-12, 2014

13 Lee JH, "Intracellular reprogramming of expression, glycosylation, and function of a plant-derived antiviral therapeutic monoclonal antibody" 8 : e68772-, 2013

14 Bakker A, "First administration to humans of a monoclonal antibody cocktail against rabies virus: safety, tolerability, and neutralizing activity" 26 : 5922-5927, 2008

15 Wang DD, "Expression, purification and characterization of a human single-chain Fv antibody fragment fused with the Fc of an IgG1 targeting a rabies antigen in Pichia pastoris" 86 : 75-81, 2012

16 Kim Y, "Expression of human interferon alpha-1 with enhanced stability via the tagging system of a stabilizing peptide" 63 : 140-146, 2009

17 Turki I, "Engineering of a recombinant trivalent single-chain variable fragment antibody directed against rabies virus glycoprotein G with improved neutralizing potency" 57 : 66-73, 2014

18 Myun PS, "Effects of novel peptides derived from the acidic tail of synuclein (ATS) on the aggregation and stability of fusion proteins" 17 : 251-260, 2004

19 Sikes RK, "Effective protection of monkeys against death from street virus by post-exposure administration of tissueculture rabies vaccine" 45 : 1-11, 1971

20 Safar F, "Development trends for generation of single-chain antibody fragments" 36 : 1-12, 2014

21 Todorovska A, "Design and application of diabodies, triabodies and tetrabodies for cancer targeting" 248 : 47-66, 2001

22 Richard F, "Current and future tools for global canine rabies elimination" 100 : 220-225, 2013

23 Jaap G, "Comparison of an anti-rabies human monoclonal antibody combination with human polyclonal anti-rabies immune globulin" 193 : 796-801, 2006

24 Liu X, "Characterization of a human antibody fragment Fab and its calcium phosphate nanoparticles that inhibit rabies virus infection with vaccine" 6 : e19848-, 2011

25 Weisser NE, "Applications of single-chain variable fragment antibodies in therapeutics and diagnostics" 27 : 502-520, 2009

26 Nelson AL, "Antibody fragments: hope and hype" 2 : 77-83, 2010

27 Wu X, "Affinity maturation of a human single-chain Fv by phage display" 21 : 156-159, 2001

28 Reed LJ, "A simple method of estimating fifty per cent endpoints" 27 : 493-497, 1938

29 Duan Y, "A novel disulfide-stabilized single-chain variable antibody fragment against rabies virus G protein with enhanced in vivo neutralizing potency" 51 : 188-196, 2012

30 Cheng Y, "A VL-linker-VH orientation dependent single chain variable antibody fragment against rabies virus G protein with enhanced neutralizing potency in vivo" 23 : 24-32, 2015