Nucleocapsid protein of cell culture-adapted Seoul virus strain 80-39: analysis of its encoding sequence, expression in yeast and immuno-reactivity.
Seoul virus (SEOV) is a hantavirus causing a mild to moderate form of hemorrhagic fever with renal syndrome that is distributed mainly in Asia. The nucleocapsid (N) protein-encoding sequence of SEOV (strain 80-39) was RT-PCR-amplified and cloned into a yeast expression vector containing a galactose-inducible promoter. A survey of the pattern of synonymous codon preferences for a total of 22 N protein-encoding hantavirus genes including 13 of SEOV strains revealed that there is minor variation in codon usage by the same gene in different viral genomes. Introduction of the expression plasmid into yeast Saccharomyces cerevisiae resulted in the high-level expression of a hexahistidine-tagged N protein derivative. The nickel-chelation chromatography purified, yeast-expressed SEOV N protein reacted in the immunoblot with a SEOV-specific monoclonal antibody and certain HTNV- and PUUV-cross-reactive monoclonal antibodies. The immunization of a rabbit with the recombinant N protein resulted in the induction of a high-titered antibody response. In ELISA studies, the N protein was able to detect antibodies in sera of experimentally infected laboratory rats and in human anti-hantavirus-positive sera or serum pools of patients from different geographical origin. The yeast-expressed SEOV N protein represents a promising antigen for development of diagnostic tools in serology, sero prevalence studies and vaccine development.
Schmidt
Jonas
J
Institute of Virology, Charité Medical School, Campus Mitte, 10098 Berlin, Germany.
Jandrig
Burkhard
B
Klempa
Boris
B
Yoshimatsu
Kumiko
K
Arikawa
Jiro
J
Meisel
Helga
H
Niedrig
Matthias
M
Pitra
Christian
C
Krüger
Detlev H
DH
Ulrich
Rainer
R
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000914:Antibodies, Viral / Q000097:blood; D003001:Cloning, Molecular; D003062:Codon / Q000235:genetics; D003429:Cross Reactions; D018076:DNA, Complementary / Q000737:chemistry / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005814:Genes, Viral; D006480:Hemorrhagic Fever with Renal Syndrome / Q000175:diagnosis / Q000453:epidemiology / Q000276:immunology / Q000821:virology; D006801:Humans; D019590:Nucleocapsid Proteins / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D010802:Phylogeny; D011110:Polymorphism, Genetic; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029263:Seoul virus / Q000235:genetics*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D003062:Codon; D018076:DNA, Complementary; D019590:Nucleocapsid Proteins; D012367:RNA, Viral
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