Immunogenic and antigenic properties of recombinant soluble glycoprotein of rabies virus.
Rabies virus glycoprotein is a type I transmembrane protein exposed on the surface on the mature virus particle that induces virus neutralizing antibodies. In the present study, 60 amino acid C-terminal hydrophobic anchor (transmembrane) and cytoplasmic domains of glycoprotein were deleted from full-length glycoprotein and fused with polyhistidine tag. The N-terminal viral signal peptide was also replaced with CD33 signal peptide for efficient secretion in mammalian cells. Following transfection of Madin Darby bovine kidney (MDBK) cells with plasmid encoding this soluble form of glycoprotein, polyclonal populations of stably transfected resistant cells were obtained after G418 selection. The protein was expressed as a glycosylated protein and secreted outside the cells utilizing N-terminal CD33 signal peptide. The secreted soluble glycoprotein was purified from cell culture supernatant by Ni--agarose affinity chromatography utilizing C-terminal polyhistidine tag. Like full-length glycoprotein, the expressed recombinant soluble glycoprotein was found to be immunogenic when injected in rabbits. In this study, we have assessed the potential of recombinant soluble glycoprotein as diagnostic antigen in ELISA and found that this recombinant protein can be used as diagnostic antigen in ELISA for detecting anti-glycoprotein antibodies in immunized host.
Gupta
Praveen K
PK
National Biotechnology Centre, Indian Veterinary Research Institute, Izatnagar, 243122 UP, India. praveenkg_brly@sancharnet.in
Sharma
Sameer
S
Walunj
Sameer S
SS
Chaturvedi
V K
VK
Raut
Ashwin A
AA
Patial
Sonika
S
Rai
A
A
Pandey
K D
KD
Saini
M
M
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology*; D015153:Blotting, Western; D002846:Chromatography, Affinity; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics; D004283:Dog Diseases / Q000821:virology; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D006023:Glycoproteins / Q000235:genetics / Q000276:immunology*; D007114:Immunization; D016133:Polymerase Chain Reaction; D011817:Rabbits; D011818:Rabies / Q000821:virology; D011820:Rabies virus / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity; D014162:Transfection; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology*
D000914:Antibodies, Viral; D000956:Antigens, Viral; D004279:DNA, Viral; D006023:Glycoproteins; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; C064951:glycoprotein G, Rabies virus