Cloning, expression and purification of the coat protein of encephalitis virus (DIEV) infecting Dicentrarchus labrax.
The coat protein gene from encephalitis virus infecting Dicentrarhus labrax (DIEV) has been cloned by gene amplification, sequenced and expressed in Escherichia coli. DNA sequencing has revealed an open reading frame of 1017 bases encoding a polypeptide of 338 amino acids. The sequence similarities between the DIEV coat protein gene and the same gene in five encephalitis viruses infected other fish species were over 71.5% at the nucleotide level and over 79.5% at the amino acid level. These results indicate that the nodaviruses that cause encephalopathy and retinopathy in fishes are very closed related. E. coli cells harbouring the plasmid containing the DIEV gene can produce the viral coat protein. An efficient purification scheme using a Sepharore-Ni+2 column is presented. This, gives approx. 10 mg of more than 95% pure protein per gr of E. coli culture.
Sideris
D C
DC
University of Athens, Dept. of Biochemistry - Molecular biology, Greece.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001492:Bass / Q000821:virology*; D002213:Capsid / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D036022:Capsid Proteins; D002846:Chromatography, Affinity / Q000379:methods; D003001:Cloning, Molecular; D004668:Encephalitis Viruses / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D012328:RNA Viruses / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid
D036022:Capsid Proteins; C108344:Diev coat protein, Dicentrarchus labrax encephalitis virus; D011994:Recombinant Proteins