Temporal nature of the promoter and not relative strength determines the expression of an extensively processed protein in a baculovirus system.
We demonstrate that the expression of extensively modified and secreted heterologous proteins synthesized in the baculovirus expression vector system (BEVS) depends on the temporal nature of the promoter transcribing the foreign gene. The beta subunit of the human chorionic gonadotropin, an extensively modified secretory glycoprotein hormone was expressed under the transcriptional control of the AcNPV basic protein gene promoter (MP) and the polyhedrin gene promoter (POL), respectively. MP, activated late in the infection cycle, is a weaker promoter when compared to the stronger very late POL promoter. Levels of secretion, immunoreactivity and bioactivity of recombinant proteins, beta hCGMP and beta hCGPOL synthesized under control of the MP and POL promoter were compared. Secretion of beta h CGMP was relatively higher. Enzymatic analysis revealed that the synthesized protein was sialylated. Receptor binding assays and testosterone stimulation assays in a mouse Leydig cell system demonstrated that on a unit protein basis, beta hCGMP was biologically more active than beta hCGPOL.
Sridhar
P
P
Eukaryotic Gene Expression, National Institute of Immunology, New Delhi, India.
Panda
A K
AK
Pal
R
R
Talwar
G P
GP
Hasnain
S E
SE
D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D002478:Cells, Cultured; D006063:Chorionic Gonadotropin / Q000235:genetics* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D007313:Insecta; D000077183:Occlusion Body Matrix Proteins; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D014764:Viral Proteins / Q000235:genetics*; D015678:Viral Structural Proteins
D006063:Chorionic Gonadotropin; D000077183:Occlusion Body Matrix Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins; D015678:Viral Structural Proteins; C009829:polyhedrin protein, Nucleopolyhedrovirus