Figure 3
From: Refined study of the interaction between HIV-1 p6 late domain and ALIX
Y2H-TPCR screening assay used to map the HIV-1 p6 ΔSQKQ -binding site in ALIX. Random tagged PCR was performed using full length AIP-1 DNA sequence as a template according to a previously described technique {Chen, 2005 #33}. The resulting library of AIP-1 fragments was amplified in Escherichia coli DH5α. The ALIX library was cotransformed with pGBKT7-p6ΔSQKQ bait into AH109 yeast and streaked onto SD/-Ade/-His/-Leu/-Trp plates. Clones growing on selective plates after 4–5 days at 30°C were recovered by transformation into bacteria, and inserts were sequenced. The amino acid sequence of ALIX (aa: 391–510, REFSEQ: accession NM_013374.3) that is represented, corresponds to the p6ΔSQKQ-binding fragment identified in this work.Inset: p6, p6ΔSQKQ, p6 Y36A and p9 were tested for interaction with ALIX391–510 in experimental conditions similar to those described in Figure 2B.