Baculovirus expression and affinity purification of protein E2 of classical swine fever virus strain Alfort/187.
The genome region encoding the major envelope glycoprotein E2 (gp55) of the classical swine fever virus (CSFV) strain Alfort/187 was cloned and sequenced. The E2 gene, either with or without additional authentic 5'-terminal sequences coding for two variants of a putative signal sequence, was used to construct recombinant baculoviruses expressing the respective glycosylated and nonglycosylated E2 protein in insect cells. The signal sequences mediated glycosylation in insect cells, but no efficient secretion of the protein into the cell culture supernatant was observed. Six histidine residues introduced at the carboxy terminus of E2 allowed purification of E2 protein by Ni(2+)-chelate affinity chromatography. The proteins obtained were characterized and their immunological properties were compared by western blot analysis.
Ruggli
N
N
Institute of Virology and Immunoprophylaxis, Mittelhäusern, Switzerland.
Moser
C
C
Mitchell
D
D
Hofmann
M
M
Tratschin
J D
JD
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004279:DNA, Viral; D005814:Genes, Viral; D005822:Genetic Vectors; D006023:Glycoproteins / Q000378:metabolism; D008969:Molecular Sequence Data; D017924:Nucleopolyhedroviruses / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D012995:Solubility; D013552:Swine; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification*
D004279:DNA, Viral; D006023:Glycoproteins; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C087086:glycoprotein E2, classical swine fever virus