Analysis of cross reactivity of retrovirus proteases using a vaccinia virus-T7 RNA polymerase-based expression system.
We have used the vaccinia virus-T7 RNA polymerase-based expression system for studies on the activity of proteases from various retroviruses on homologous and heterologous Gag polyproteins in eukaryotic cells. Proteases from human immunodeficiency virus (HIV) types 1 and 2, equine infectious anaemia virus, human T cell leukaemia virus type 1 and human spumavirus were produced and were shown to cleave their cognate Gag substrates produced in trans. Analysis of cross reactivity revealed that lentivirus proteases cleaved only lentivirus Gag proteins and oncovirus proteases acted primarily on oncovirus Gag proteins. The HIV-2 protease cleaved the HIV-1 Gag precursor almost as efficiently as HIV-1 protease. Expression of the 5' end of the human spumavirus pol gene revealed that it encodes a functional protease that acts specifically on the human spumavirus Gag polyprotein. This assay will allow further investigation on the activity and specificity of retrovirus proteases in eukaryotic cells.
Luukkonen
B G
BG
Microbiology and Tumorbiology Center, Karolinska Institute, Stockholm, Sweden.
Tan
W
W
Fenyö
E M
EM
Schwartz
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D015683:Gene Products, gag / Q000378:metabolism*; D015684:Gene Products, pol; D005822:Genetic Vectors; D006678:HIV / Q000201:enzymology / Q000235:genetics; D006367:HeLa Cells; D015368:Human T-lymphotropic virus 1 / Q000201:enzymology / Q000235:genetics; D006801:Humans; D004860:Infectious Anemia Virus, Equine / Q000201:enzymology / Q000235:genetics; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012190:Retroviridae / Q000201:enzymology* / Q000235:genetics; D016092:Spumavirus / Q000201:enzymology / Q000235:genetics; D013379:Substrate Specificity; D014616:Vaccinia virus / Q000235:genetics*; D014764:Viral Proteins
D015683:Gene Products, gag; D015684:Gene Products, pol; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D010450:Endopeptidases