FIG. 4.

Characterization of the Vpr-LysRS interaction in vivo. (A) Amino acid sequence of wild-type Vpr with previously defined functional domains shown (47, 50, 71). N- and C-terminal Vpr deletion mutants were cloned into PV44ER.Lex and expressed in yeast. Broken lines represent the region of protein expressed by the mutants, and numbers indicate the amino acid positions. (B) Vpr deletion mutants were transformed into L40 along with GalAd-c2.10 and grown on His⁻ plates containing 0 to 50 mM 3-AT to measure the level of transcription from the HIS3 gene. Yeast strains expressing strongly interacting proteins remain prototrophic for histidine in the presence of high concentrations of 3-AT. The assay was carried out on three separate transformants for each Vpr mutant. (C) The β-galactosidase activity of yeast strains expressing Vpr hybrid proteins along with GalAD-c2.10 was measured in a chemiluminescence assay. The background was determined by L40 expressing LexA-Vprwt and was around 200 RLU. Values are means of triplicate assays performed on four independent transformants.