Abstract
Yellow head virus (YHV) is an invertebrate nidovirus that is highly pathogenic for marine shrimp. Nucleotide sequence analysis indicated that the YHV ORF2 gene encodes a basic protein (pl = 9.9) of 146 amino acids with a predicted molecular weight of 16,325.5 Da. The deduced amino acid sequence indicated a predominance of basic (15.1%), acidic (9.6%) and hydrophilic polar (34.3%) residues and a high proportion proline and glycine residues (16.4%). The ORF2 gene was cloned and expressed in Escherichia coli as a M-r = 21 kDa His(6)-Protein that reacted with YHV nucleoprotein (p20) monoclonal antibody. Segments representing the four linear quadrants of the nucleoprotein were also expressed in E. coli as GST-fusion proteins. Immunoblot analysis using YHV polyclonal rabbit antiserum indicated the presence of linear epitopes in all except the V-37-Q(74) quadrant. Immunoblot analysis of the GST-fusion proteins and C-terminally truncated segments of the nucleoprotein allowed mapping of YHV monoclonal antibodies Y19, Y20 and YII4 to linear epitopes in the acidic domain between amino acids I-116 and E-137. The full-length nucleoprotein was expressed at high level in E. coli and was easily Purified in quantity from the Soluble cell fraction by Ni+-NTA affinity chromatography. (c) 2005 Elsevier B.V. All rights reserved.
