[Expression and purification of recombinant SARS coronavirus spike protein].
A novel coronavirus (SARS-coronavirus, SARS-CoV) was discovered in association with cases of severe acute respiratory syndrome (SARS) recently. The first step in coronavirus infection is binding of the viral spike protein to certain receptor on host cells. The spike protein is the main surface antigen of the coronavirus and there should be antibodies against spike protein in patients serum. Thus, to develop and expression protein fragment from spike protein gene are the purposes of this experiment. Partial spike gene fragments (751-1925 bp, 2005-3410 bp, 1-1925 bp and 32-3659 bp) and its intact gene were cloned into pET32 or pGEX vectors, and transformed into competent Escherichia coli BL21(DE3) (pLysS), respectively. 63, 78, 98, 160 and 164 kD fusion proteins were successfully expressed with amounts of 35%, 34%, 24%, 17% and 5% of total cell protein. The soluble parts of the cell crude extract were then partially purified by GST affinity chromatography.
Yu
Hao
H
Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, Shanghai 200032, China.
Yang
Yong
Y
Zhang
Wei
W
Xie
You-Hua
YH
Qin
Jun
J
Wang
Yuan
Y
Zheng
Hua-Bao
HB
Zhao
Guo-Ping
GP
Yang
Sheng
S
Jiang
Wei-Hong
WH
D015153:Blotting, Western; D004926:Escherichia coli / Q000235:genetics; D008562:Membrane Glycoproteins / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification; D045473:Severe acute respiratory syndrome-related coronavirus / Q000737:chemistry* / Q000276:immunology; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification
D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV