EXPRESSION AND SECRETION OF JAPANESE ENCEPHALITIS-VIRUS NONSTRUCTURAL PROTEIN-NS1 BY INSECT CELLS USING A RECOMBINANT BACULOVIRUS

被引:32
作者
FLAMAND, M [1 ]
DEUBEL, V [1 ]
GIRARD, M [1 ]
机构
[1] INST PASTEUR,UNITE VIROL MOLEC,F-75724 PARIS 15,FRANCE
关键词
D O I
10.1016/0042-6822(92)90258-Q
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
The nonstructural protein NS1 of Japanese encephalitis virus (JEV) was expressed at a high level under the control of the polyhedrin promoter in Spodoptera frugiperda (Sf9) insect cells using a recombinant baculovirus. Recombinant NS1 was designed to contain its natural signal sequence at its N-terminus and no C-terminal hydrophobic domain that could act as a membrane anchor. This recombinant protein exhibited similar size to native NS1 expressed in Aedes albopictus (C6/36) insect cells infected with wild-type JEV. The signal sequence of NS1 allowed translocation of the protein into the endoplasmic reticulum where it underwent glycosylation. A small fraction of synthesized NS1 was able, in the absence of any other viral protein, to associate as a homodimer, showing similar characteristics to the native dimer. Interestingly, this recombinant dimeric form seemed to be exported and released in the extracellular medium of infected cell culture. During its transport, one of the two Winked oligosaccharides of the polymannose type was processed to an endoglycosidase H-resistant form, suggesting that the protein had passed through the Golgi compartment before reaching the cell surface. Moreover, Triton X-114 partitioning analysis showed that monomeric NS1 behaved essentially as a hydrophilic protein, whereas both intracellular and extracellular dimeric NS1 were either free of or associated to membraneous components. © 1992.
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页码:826 / 836
页数:11
相关论文
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[1]  
BORDIER C, 1981, J BIOL CHEM, V256, P1604
[2]   DENGUE VIRIONS AND ANTIGENS IN BRAIN AND SERUM OF INFECTED MICE [J].
BRANDT, WE ;
CARDIFF, RD ;
RUSSELL, PK .
JOURNAL OF VIROLOGY, 1970, 6 (04) :500-&
[3]  
Brinton MA., 1986, TOGAVIRIDAE FLAVIVIR, P327
[4]   MOLECULAR SIZE AND CHARGE RELATIONSHIPS OF SOLUBLE COMPLEMENT-FIXING ANTIGENS OF DENGUE VIRUSES [J].
CARDIFF, RD ;
MCCLOUD, TG ;
BRANDT, WE ;
RUSSELL, PK .
VIROLOGY, 1970, 41 (03) :569-&
[5]   FLAVIVIRUS GENOME ORGANIZATION, EXPRESSION, AND REPLICATION [J].
CHAMBERS, TJ ;
HAHN, CS ;
GALLER, R ;
RICE, CM .
ANNUAL REVIEW OF MICROBIOLOGY, 1990, 44 :649-688
[6]   PURIFICATION AND ANALYSIS OF INFECTIOUS VIRIONS AND NATIVE NONSTRUCTURAL ANTIGENS FROM CELLS INFECTED WITH TICK-BORNE ENCEPHALITIS-VIRUS [J].
CROOKS, AJ ;
LEE, JM ;
DOWSETT, AB ;
STEPHENSON, JR .
JOURNAL OF CHROMATOGRAPHY, 1990, 502 (01) :59-68
[7]  
DEPSRES P, 1988, ANN INST PASTEUR VIR, V139, P59
[8]   CHARACTERIZATION OF YELLOW-FEVER VIRUS PROTEINS-E AND NS1 EXPRESSED IN VERO AND SPODOPTERA-FRUGIPERDA CELLS [J].
DESPRES, P ;
GIRARD, M ;
BOULOY, M .
JOURNAL OF GENERAL VIROLOGY, 1991, 72 :1331-1342
[9]   VIRION AND SOLUBLE-ANTIGENS OF JAPANESE ENCEPHALITIS-VIRUS [J].
ECKELS, KH ;
HETRICK, FM ;
RUSSELL, PK .
INFECTION AND IMMUNITY, 1975, 11 (05) :1053-1060
[10]   PROPER PROCESSING OF DENGUE VIRUS NONSTRUCTURAL GLYCOPROTEIN-NS1 REQUIRES THE N-TERMINAL HYDROPHOBIC SIGNAL SEQUENCE AND THE DOWNSTREAM NONSTRUCTURAL PROTEIN-NS2A [J].
FALGOUT, B ;
CHANOCK, R ;
LAI, CJ .
JOURNAL OF VIROLOGY, 1989, 63 (05) :1852-1860