1. Efficient copurification of selected co-opted host proteins with the viral replicase from yeast coexpressing TPR-domain protein or cyclophilin A (CypA).
Efficient copurification of selected co-opted host proteins with the viral replicase from yeast coexpressing TPR-domain protein or cyclophilin A (CypA). The viral replicase was purified through FLAG-tagged p33 from yeast extracts using a FLAG-affinity column. (A to D) For the top panel, Western blot analysis of copurified HA-tagged host protein expressed from the chromosomal location with anti-HA antibody was performed. For the middle panel, Western blot analysis of the purified Flag-p33 using anti-FLAG antibody was performed. For the bottom panel, Western blot analysis of total protein extract with anti-HA antibody was used to detect the total amount of the HA-tagged host protein expressed from the chromosomal location in each sample. Lane 1, negative control based on yeast expressing His6-tagged p33 and the HA-tagged host protein; lane 2, yeast coexpressing Flag-p33 and the HA-tagged host protein; lane 3, yeast coexpressing Flag-p33 and the HA-tagged host protein in combination with TPR-domain protein; lane 4, yeast coexpressing Flag-p33 and the HA-tagged host protein in combination with CypA protein. All yeast strains actively replicate the TBSV repRNA (data not shown). Each experiment was repeated. (E) Western blot analysis of total protein extracts from yeasts expressing the shown proteins through anti-His antibody.
Daniel Barajas et al. J. Virol. 2015;89: