Production and characterization of a recombinant single-chain antibody against Hantaan virus envelop glycoprotein.
Hantaan virus (HTNV) is the type of Hantavirus causing hemorrhagic fever with renal syndrome, for which no specific therapeutics are available so far. Cell type-specific internalizing antibodies can be used to deliver therapeutics intracellularly to target cell and thus, have potential application in anti-HTNV infection. To achieve intracellular delivery of therapeutics, it is necessary to obtain antibodies that demonstrate sufficient cell type-specific binding, internalizing, and desired cellular trafficking. Here, we describe the prokaryotic expression, affinity purification, and functional testing of a single-chain Fv antibody fragment (scFv) against HTNV envelop glycoprotein (GP), an HTNV-specific antigen normally located on the membranes of HTNV-infected cells. This HTNV GP-targeting antibody, scFv3G1, was produced in the cytoplasm of Escherichia coli cells as a soluble protein and was purified by immobilized metal affinity chromatography. The purified scFv possessed a high specific antigen-binding activity to HTNV GP and HTNV-infected Vero E6 cells and could be internalized into HTNV-infected cells probably through the clathrin-dependent endocytosis pathways similar to that observed with transferrin. Our results showed that the E. coli-produced scFv had potential applications in targeted and intracellular delivery of therapeutics against HTNV infections.
Yang
Jie
J
Department of Nephrology, Tangdu Hospital, Fourth Military Medical University, Xi'an, Shaanxi, People's Republic of China.
Chen
Rui
R
Wei
Junxia
J
Zhang
Fanglin
F
Zhang
Yong
Y
Jia
Lintao
L
Yan
Yan
Y
Luo
Wen
W
Cao
Yunxin
Y
Yao
Libo
L
Sun
Jifeng
J
Xu
Zhikai
Z
Yang
Angang
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002522:Chlorocebus aethiops; D002846:Chromatography, Affinity; D004705:Endocytosis; D004926:Escherichia coli / Q000235:genetics; D018063:Hantaan virus / Q000276:immunology*; D008969:Molecular Sequence Data; D011485:Protein Binding; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D017422:Sequence Analysis, DNA; D057127:Single-Chain Antibodies / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000276:immunology*
D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; D014759:Viral Envelope Proteins