FIGURE 2.

Dug3p has functional GATaseII domain. A, multiple sequence alignment of GATaseII domain of Dug3p with the GATaseII domain of glutamine phosphoribosyl amidotransferase (PurF), glucosamine-6-phosphate synthase (Glms), and asparagine synthetase (Asn). Multiple sequence alignment was carried out by the ClustalW program. Residues predicted to be important for activity are highlighted and indicated by a bold arrow. Ec, E. coli; Sc, S. cerevisiae. B, DUG3 wild type and mutant constructs in p416TEF vector were transformed into met15Δ dug3Δ strains of S. cerevisiae, and transformants were grown in minimal supplemented medium. Cells were harvested, washed, and resuspended to an A₆₀₀ = 2. Serial dilutions of A₆₀₀ = 0.2, 0.02, 0.002, and 0.0002 were made, and 5 μl of each dilution was spotted on minimal medium having methionine or glutathione as sulfur source. Plates were incubated at 30 °C for 3 days.