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KMID : 0364820100460020140
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Korean Journal of Microbiology
2010 Volume.46 No. 2 p.140 ~ p.147
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Production of the Polyclonal Antibody That Recognizes the Mutant M Protein of Japanese Encephalitis Virus: Role of Its Charged Residues in Virus Production
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Kim Jung-Min
Yun Sang-Im
Song Byung-Hak
Kim Jin-Kyung
Lee Young-Min
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Abstract
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Japanese encephalitis virus (JEV), a member of the mosquito-borne flaviviruses, causes epidemics of viral encephalitis in the Southeastern Asia. JEV is a small enveloped virus with a positive-sense RNA genome; the infectious virion consists of three structural proteins, namely capsid, membrane (M; a mature form of its prM precursor), and envelope proteins. Here, we investigated a role of the charged residues found at the N-terminus of the JEV M protein in virus production. Using an infectious JEV cDNA, we generated two mutant cDNAs, Mm1 and Mm2, by charged-to-alanine substitution for E9 and K15K16E17 residues of the M protein, respectively. By transfection of wild-type or each of the two mutant RNAs transcribed from the corresponding cDNAs, we found that Mm2, but not Mm1, had a ~3-log decrease in virus production, even though a comparable amount of all three structural proteins were produced in transfected cells. Interestingly, the prM protein expressed in Mm2 RNA-transfected cells was not recognized by the polyclonal antiserum raised against the N-terminal 44 amino acids of the wild type M protein, but reacted to the antiserum raised against the corresponding region of the mutant Mm2. Our results indicate that three charged residues (K15K16E17) in JEV M protein play a role in virus production. Two polyclonal antisera specifically recognizing the wild-type or Mm2 version of the M protein would provide a useful reagent for the functional study of this protein in the virus life cycle.
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KEYWORD
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Flavivirus, Japanese encephalitis virus, M protein, prM protein
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