Fig. 3. Schematic representation of the split-ubiquitin assay. Preparation of the constructs is described in materials and methods. Cub-PLV was linked to the C-terminal of Rer1p. NubG or NubI (not shown) was linked to the N-terminal of Mns1p, Sec12p, and Alg5p, and to the C-terminal of Ost1p. Interaction between NubG-Mns1p or NubG-Sec12p and Rer1p-Cub-PLV results in the association of NubG with Cub leading to cleavage of PLV in the cytosol and a positive ß-galactosidase assay. In contrast, the lack of interaction between Ost1p-NubG and Rer1p-Cub-PLV gives a negative ß-galactosidase assay. The topology of NubG-Alg5p is also shown. The lengths of the protein domains are not to scale.
