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. 2006 Jun 15;20(12):1609–1620. doi: 10.1101/gad.385706

Figure 2.

Figure 2.

Interaction of Eno2p with tRK1. (A) Detection of the interaction between Eno1/2p and tRK1 in vivo by the three-hybrid approach. The strategy is presented in the upper panel. MS2 RNA-fused tRK1 or tRK2 (nonimported RNA) were assayed for interaction with either Eno1p or Eno2p. Empty MS2 RNA gene-containing vector (MS2) and empty pGAD-GH plasmid (pGAD) were used as controls. To detect interaction, yeast L40-coat strain transfected cells were grown in the absence or presence of 10 mM 3AT, as indicated. (B–D) Analysis of RNA–protein interactions by gel-shift. Autoradiographs of native PAGE-separations of labeled tRK1 in the presence of preMsk1p (B) and/or Eno2p (C). Deacylated tRK1 [tRK1(da)] and aminoacylated tRK2 [tRK2(aa)] were used as controls of the specificity of interaction. Concentrations of the recombinant proteins in nM are indicated below the panels. (KMC) tRK1–preMsk1 complex; (KEC) tRK1–Eno2p complex; (tRK1) unbound labeled tRNA. (D) The effect of Eno2p presence on tRK1–preMsk1p complex formation.