Table IV

Pex5p has distinct binding sites for the C3HC4 RING finger domain of Pex12p and PTS1 proteins.

Yeast cells (strain SFY526), transformed with plasmids coding for a Gal4p DNA-binding domain-PTS1 fusion protein, a Gal4p activation domain-Pex5p fusion protein, and a third (conditionally expressed from the Met25 promotor) HA-tagged fusion protein (PTS1 or Pex12p(275–359), were grown for 72 h in minimal dropout medium without leucine and tryptophan, in the absence (−) or presence (+) or 1 nm methionine. The cells were assayed for β-galactosidase using o-nitrophenyl-β-d-galactopyranoside as the substrate (see “Experimental Procedures”). The optical densities were measured at 420 nm and normalized for culture densities (optical density at 600 nm = 10) and time (24 h). The values given are the mean (± the S.D.) of three measurements performed on each of four separate transformants. Notice that the HA-tagged fusion protein is repressed in the presence of 1 mm methionine and expressed in the absence of methionine.

Optical density
+ Methionine− Methionine
HA-PTS1600 ± 280120 ± 22
HA-Pex12p(275–359)532 ± 165596 ± 223