Proteomic analysis of shrimp white spot syndrome viral proteins and characterization of a novel envelope protein VP466.
White spot syndrome virus (WSSV) is at present one of the major pathogens in shrimp culture worldwide. The complete genome of this virus has been sequenced recently. To identify the structural and functional proteins of WSSV, the purified virions were separated by SDS-PAGE. Twenty-four protein bands were excised, in-gel digested with trypsin, and subjected to matrix-assisted laser desorption ionization-time of flight mass spectrometry and electrospray ionization tandem mass spectrometry, respectively. Eighteen proteins matching the open reading frames of WSSV genome were identified. Except for three known structural proteins and collagen, the functions of the remaining 14 proteins were unknown. Temporal analysis revealed that all the genes were transcribed in the late stage of WSSV infection except for vp121. Of the newly identified proteins, VP466 (derived from band 16) was further characterized. The cDNA encoding VP466 was expressed in Escherichia coli as a glutathione S-transferase (GST) fusion protein. Specific antibody was generated with the purified GST-VP466 fusion protein. Western blot showed that the mouse anti-GST-VP466 antibody bound specifically to a 51-kDa protein of WSSV. Immunogold labeling revealed that VP466 protein is a component of the viral envelope. Results in this investigation thus proved the effectiveness of proteomic approaches for discovering new proteins of WSSV.
Huang
Canhua
C
Department of Biological Sciences and Tropical Marine Science Institute, National University of Singapore, Singapore 119260.
Zhang
Xiaobo
X
Lin
Qingsong
Q
Xu
Xun
X
Hu
Zhihong
Z
Hew
Choy-L
CL
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D004267:DNA Viruses / Q000737:chemistry* / Q000235:genetics* / Q000472:pathogenicity / Q000648:ultrastructure; D004279:DNA, Viral / Q000235:genetics; D033364:Decapoda / Q000821:virology; D005814:Genes, Viral; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D020543:Proteome / Q000235:genetics* / Q000302:isolation & purification*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000302:isolation & purification*
D004279:DNA, Viral; D020543:Proteome; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins