Characterization of the protease of a fish retrovirus, walleye dermal sarcoma virus.
Three fish retroviruses infecting walleyes constitute the recently recognized genus called epsilonretrovirus. The founding member of this group, walleye dermal sarcoma virus (WDSV), induces benign skin tumors in the infected fish and replicates near 4 degrees C. While the viral genomic sequence is known, biochemical characterization of the virus has been limited to the identification of the mature structural and envelope proteins present in virions. We undertook this study to determine the cleavage sites in the WDSV Pro and Pol proteins and to characterize the viral protease (PR) in vitro. A recombinant PR was expressed in and purified from Escherichia coli as a larger fusion with additional nucleocapsid and reverse transcriptase residues flanking the PR domain. Autocleavage produced a functional, mature PR. Autocleavage as well as cleavage of peptides and of Gag protein by the mature PR occurred at a pH optimum of 7.0, higher than that of other retroviral proteases. Analysis of the cleavage sites identified a glutamine residue in the P2 position of all WDSV sites, both in Gag and in Pol. Amino acid sequence alignments of Gag-Pro-Pol from WDSV, walleye epidermal hyperplasia virus type 1, and walleye epidermal hyperplasia virus type 2 showed the P2 glutamine to be conserved in all cleavage sites in these three viruses. Such conservation is unprecedented in other retroviruses.
Fodor
Sharon K
SK
Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA.
Vogt
Volker M
VM
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D010450:Endopeptidases / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D030283:Epsilonretrovirus / Q000201:enzymology* / Q000235:genetics*; D008969:Molecular Sequence Data; D020815:Polyproteins / Q000378:metabolism; D011498:Protein Precursors / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
D020815:Polyproteins; D011498:Protein Precursors; D011994:Recombinant Proteins; D010450:Endopeptidases