Antigenicity and receptor-binding ability of recombinant SARS coronavirus spike protein.
Severe acute respiratory syndrome (SARS) is an emerging infectious disease associated with a novel coronavirus and causing worldwide outbreaks. SARS coronavirus (SARS-CoV) is an enveloped RNA virus, which contains several structural proteins. Among these proteins, spike (S) protein is responsible for binding to specific cellular receptors and is a major antigenic determinant, which induces neutralizing antibody. In order to analyze the antigenicity and receptor-binding ability of SARS-CoV S protein, we expressed the S protein in Escherichia coli using a pET expression vector. After the isopropyl-beta-D-thiogalactoside induction, S protein was expressed in the soluble form and purified by nickel-affinity chromatography to homogeneity. The amount of S protein recovered was 0.2-0.3mg/100ml bacterial culture. The S protein was recognized by sera from SARS patients by ELISA and Western blot, which indicated that recombinant S protein retained its antigenicity. By biotinylated ELISA and Western blot using biotin-labeled S protein as the probe, we identified 130-kDa and 140-kDa proteins in Vero cells that might be the cellular receptors responsible for SARS-CoV infection. Taken together, these results suggested that recombinant S protein exhibited the antigenicity and receptor-binding ability, and it could be a good candidate for further developing SARS vaccine and anti-SARS therapy.
Ho
Tin-Yun
TY
Institute of Chinese Medical Science, China Medical University, ROC, Taichung, Taiwan
Wu
Shih-Lu
SL
Cheng
Shin-Ei
SE
Wei
Yen-Chiao
YC
Huang
Shan-Ping
SP
Hsiang
Chien-Yun
CY
D000595:Amino Acid Sequence; D000818:Animals; D000956:Antigens, Viral / Q000235:genetics; D001483:Base Sequence; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000235:genetics; D006801:Humans; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D011991:Receptors, Virus / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics / Q000276:immunology* / Q000378:metabolism* / Q000472:pathogenicity; D017386:Sequence Homology, Amino Acid; D064370:Spike Glycoprotein, Coronavirus; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*
D000956:Antigens, Viral; D004279:DNA, Viral; D008562:Membrane Glycoproteins; D011991:Receptors, Virus; D011994:Recombinant Proteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus