Deletion of the myristylation signal allows high-level production of the hepatitis B virus large surface glycoprotein preS1 with vaccinia virus recombinants.
We have investigated the requirements necessary for high-level production of the hepatitis B virus (HBV strain ayw) large surface glycoprotein preS1 with vaccinia virus (VV) recombinants. In earlier studies, only nanogram amounts of preS1 could be obtained from cells infected with an appropriate recombinant VV carrying the preS1 gene under the transcriptional control of a conventional VV promoter (p7.5). Here, we report that the use of an improved promoter system, i.e., the bacteriophage T7 polymerase/VV hybrid expression system (T7/EMC system) in combination with a G-C conversion at position 5 of the preS1 open reading frame, deleting the myristylation motif of the polypeptide, results in an at least 12-fold increase in preS1 expression compared to the wild-type preS1 expressed with the strongest homologous VV promoter system known so far. Although the T7/EMC promoter system was most effective, improved expression of the modified preS1 (preS1dMyr) is independent from the promoter system used, from the insertion locus of the modified preS1 within the VV genome and also from the cell line used for expression studies.
Pfleiderer
M
M
IMMUNO AG Biomedical Research Center, Austria. pfmi@immuno.co.at
Falkner
F G
FG
Dorner
F
F
D000222:Adaptation, Physiological; D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D005785:Gene Conversion; D015967:Gene Expression Regulation, Viral; D005822:Genetic Vectors; D006514:Hepatitis B Surface Antigens / Q000235:genetics*; D006515:Hepatitis B virus / Q000235:genetics*; D006801:Humans; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism*; D016366:Open Reading Frames; D011498:Protein Precursors / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics; D014616:Vaccinia virus / Q000235:genetics*; D014709:Vero Cells
D006514:Hepatitis B Surface Antigens; D009227:Myristic Acids; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C050806:presurface protein 1, hepatitis B surface antigen; D019814:Myristic Acid