Generation of human single-chain variable fragment antibodies specific to dengue virus non-structural protein 1 that interfere with the virus infectious cycle.
Severe forms of dengue virus (DENV) infection frequently cause high case fatality rate. Currently, there is no effective vaccine against the infection. Clinical cases are given only palliative treatment as specific anti-DENV immunotherapy is not available and it is urgently required. In this study, human single-chain variable fragment (HuScFv) antibodies that bound specifically to the conserved non-structural protein-1 (NS1) of DENV and interfered with the virus replication cycle were produced by using phage display technology. Recombinant NS1 (rNS1) of DENV serotype 2 (DENV2) was used as antigen in phage bio-panning to select phage clones that displayed HuScFv from antibody phage display library. HuScFv from two phagemid transformed E. coli clones, i.e., clones 11 and 13, bound to the rNS1 as well as native NS1 in both secreted and intracellular forms. Culture fluids of the HuScFv11/HuScFv13 exposed DENV2 infected cells had significant reduction of the infectious viral particles, implying that the antibody fragments affected the virus morphogenesis or release. HuScFv epitope mapping by phage mimotope searching revealed that HuScFv11 bound to amino acids 1-14 of NS1, while the HuScFv13 bound to conformational epitope at the C-terminal portion of the NS1. Although the functions of the epitopes and the molecular mechanism of the HuScFv11 and HuScFv13 require further investigations, these small antibodies have high potential for development as anti-DENV biomolecules.
Division of Molecular Medicine; Department of Research and Development; Faculty of Medicine Siriraj Hospital; Mahidol University; Bangkok, Thailand.
Ornnuthchar
O
Poungpair
Department of Biochemistry; Faculty of Sciences; Kasetsart University; Bangkok, Thailand.
Kunan
K
Bangphoomi
Division of Molecular Medicine; Department of Research and Development; Faculty of Medicine Siriraj Hospital; Mahidol University; Bangkok, Thailand.
Prapaipit
P
Chaowalit
Division of Molecular Medicine; Department of Research and Development; Faculty of Medicine Siriraj Hospital; Mahidol University; Bangkok, Thailand.
Nunghathai
N
Sawasdee
Immunology Graduate Program; Department of Immunology; Faculty of Medicine Siriraj Hospital; Mahidol University; Bangkok, Thailand.
Nichapatr
N
Saokaew
Department of Biochemistry; Faculty of Sciences; Kasetsart University; Bangkok, Thailand.
Kiattawee
K
Choowongkomon
Laboratory for Research and Technology Development; Department of Parasitology; Faculty of Medicine Siriraj Hospital; Mahidol University; Bangkok, Thailand.
Wanpen
W
Chaicumpa
Division of Molecular Medicine; Department of Research and Development; Faculty of Medicine Siriraj Hospital; Mahidol University; Bangkok, Thailand.
Pa-thai
PT
Yenchitsomanus
D000330:Aedes; D000818:Animals; D002460:Cell Line; D061505:Cell Surface Display Techniques; D003715:Dengue / Q000276:immunology / Q000378:metabolism* / Q000635:transmission; D003716:Dengue Virus / Q000472:pathogenicity / Q000502:physiology*; D018604:Epitope Mapping; D000939:Epitopes / Q000276:immunology / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D006801:Humans; D006825:Hybridomas; D007167:Immunotherapy / Q000379:methods*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism*; D057127:Single-Chain Antibodies / Q000302:isolation & purification*; D017361:Viral Nonstructural Proteins / Q000276:immunology / Q000378:metabolism*; D014771:Virion / Q000187:drug effects
D000939:Epitopes; C079340:NS1 protein, Dengue virus type 2; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; D017361:Viral Nonstructural Proteins
HuScFv; NS1; dengue virus; human single-chain variable fragment antibody; monoclonal antibody; non-structural protein-1; phage display