Purification of the extra-cellular domain of Nipah virus glycoprotein produced in Escherichia coli and possible application in diagnosis.
The glycoprotein (G) of Nipah virus (NiV) is important for virus infectivity and induction of the protective immunity. In this study, the extra-cellular domain of NiV G protein was fused with hexahistidine residues at its N-terminal end and expressed in Escherichia coli. The expression under transcriptional regulation of T7 promoter yielded insoluble protein aggregates in the form of inclusion bodies. The inclusion bodies were solubilized with 8 M urea and the protein was purified to homogeneity under denaturing conditions using nickel-nitrilotriacetic acid (Ni-NTA) affinity chromatography. The denatured protein was renatured by gradual removal of the urea. Light scattering analysis of the purified protein showed primarily monodispersity. The purified protein showed significant reactivity with the antibodies present in the sera of NiV-infected swine, as demonstrated in Western blot analysis and enzyme-linked immunosorbent assay (ELISA). Taken together, the data indicate the potential usefulness of the purified G protein for structural or functional studies and the development of immunoassay for detection of the NiV antibodies.
Eshaghi
Majid
M
Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, University Putra Malaysia, 43400 Serdang, Selangor, Malaysia.
Tan
Wen Siang
WS
Chin
Wai Kit
WK
Yusoff
Khatijah
K
D000818:Animals; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D045604:Extracellular Fluid / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000097:blood / Q000276:immunology*; D045464:Henipavirus Infections / Q000097:blood / Q000175:diagnosis / Q000276:immunology / Q000821:virology; D007118:Immunoassay / Q000379:methods*; D045405:Nipah Virus / Q000235:genetics* / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification; D013552:Swine; D014760:Viral Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification
D011993:Recombinant Fusion Proteins; D014760:Viral Fusion Proteins; D019204:GTP-Binding Proteins