Development of a rapid and convenient method for the quantification of HIV-1 budding.
In cells, the expression of Gag protein, one of the major structural proteins of retroviruses, is sufficient for budding virus-like particles (VLPs) from the cell surface. We have previously reported that spheroplasts of Saccharomyces cerevisiae expressing HIV-1 Gag proteins from an episomal plasmid constitutively released a large amount of VLPs into culture media; however, commercially available ELISA kits which detect mature capsid of HIV-1 could not detect uncleaved 55-kDa Gag proteins released from budding yeast. We therefore developed a method to quantitate VLP levels released from budding yeast by using fusion protein from HIV-1 Gag and Firefly Luciferase. This system is useful for screening cellular factor(s) involved in retrovirus budding from S. cerevisiae.
Sakuragi
Sayuri
S
Department of Viral Infections, Research Institute for Microbial Diseases, Osaka University, 3-1, Yamadaoka, Suita-City, Osaka, Japan. ssakurag@biken.osaka-u.ac.jp
Sakuragi
Jun-ichi
J
Morikawa
Yuko
Y
Shioda
Tatsuo
T
D015683:Gene Products, gag / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D015497:HIV-1 / Q000235:genetics / Q000502:physiology*; D006801:Humans; D049409:Luciferases, Firefly / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D013104:Spheroplasts / Q000235:genetics / Q000502:physiology*; D014773:Virology / Q000379:methods*; D022701:Virosomes / Q000378:metabolism
D015683:Gene Products, gag; D011993:Recombinant Fusion Proteins; D022701:Virosomes; D049409:Luciferases, Firefly