Dengue virus envelope glycoprotein can be secreted from insect cells as a fusion with the maltose-binding protein.
The maltose-binding protein (MalE) contains a signal sequence which allows its translocation in the periplasm of prokaryotic microorganisms. In this study, MalE was produced in Spodoptera frugiperda (Sf9) lepidopterian cells using the baculovirus expression system. The secretion of MalE, following cleavage of its signal sequence, to the supernatant fluid of recombinant baculovirus-infected Sf9 cells and its affinity for maltodextrin polymers allowed recovery of significant amounts (> or = 10 micrograms per 10(6) cells) of highly purified protein. The gene encoding the envelope glycoprotein E of the dengue (DEN) type 2 virus deleted of its C-terminal 102 amino acids (D2E delta 102) was fused to the MalE gene. The resulting hybrid MalE-D2E delta 102 glycoprotein was processed through the Golgi network of Sf9 cells and was secreted. It was retained on a maltodextrin column and was eluted with maltose. Antigenic and immunogenic properties dependent on the three-dimensional structure in the native E protein were preserved in the recombinant MalE-D2E delta 102 protein. Thus MalE with its signal sequence may be used as a carrier protein for production in the baculovirus system and purification of proteins which require transportation through intracellular compartments for correct folding and processing.
Staropoli
I
I
Unité des Arbovirus et Virus des Fièvres Hémorragiques, Institut Pasteur, Paris, France.
Clément
J M
JM
Frenkiel
M P
MP
Hofnung
M
M
Deubel
V
V
D000818:Animals; D000956:Antigens, Viral / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification; D002352:Carrier Proteins / Q000276:immunology*; D002460:Cell Line; D003001:Cloning, Molecular; D003715:Dengue / Q000517:prevention & control* / Q000821:virology; D003716:Dengue Virus / Q000737:chemistry / Q000276:immunology*; D058619:Maltose-Binding Proteins; D051379:Mice; D008807:Mice, Inbred BALB C; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D018411:Spodoptera / Q000166:cytology; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism
D000956:Antigens, Viral; D002352:Carrier Proteins; C051821:E-glycoprotein, Dengue virus type 2; D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins