Truncating the putative membrane association region circumvents the difficulty of expressing hepatitis C virus protein E1 in Escherichia coli.
The putative E1 of hepatitis C virus (HCV) was expressed in Escherichia coli using a glutathione-S-transferase (GST) fusion protein system. The full length E1 protein is difficult to express. A series of E1 DNA fragments was generated and used for expression vector construction. Fusion proteins containing the E1 C-terminal region could not be expressed. When this region was truncated, the fusion proteins were synthesized to high levels. The possibility of this C-terminal region hampering the production of fusion protein was further explored. A construct with this segment directly fused to the C-terminus of GST indeed generated no detectable recombinant protein. According to the predicted structure of E1, this region may have membrane-associating properties. The expression results suggest a general approach to facilitate the production of viral membrane proteins in prokaryotes. Furthermore, these recombinant E1 proteins generated as antigens were used for Western blotting with sera from HCV-infected individuals. It was found that E1 is antigenic during HCV natural infection.
Yan
B S
BS
Graduate Institute of Microbiology and Immunology, National Yang-Ming Medical College, Taiwan, ROC.
Liao
L Y
LY
Leou
K
K
Chang
Y C
YC
Syu
W J
WJ
D000956:Antigens, Viral / Q000235:genetics; D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D016174:Hepacivirus / Q000235:genetics* / Q000276:immunology; D006508:Hepatitis Antibodies / Q000097:blood; D006526:Hepatitis C / Q000276:immunology; D018937:Hepatitis C Antibodies; D018936:Hepatitis C Antigens; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology; D014773:Virology / Q000379:methods*
D000956:Antigens, Viral; D004279:DNA, Viral; C087035:E1 protein, Hepatitis C virus; D006508:Hepatitis Antibodies; D018937:Hepatitis C Antibodies; D018936:Hepatitis C Antigens; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins