Figure 4.

The N-terminal domain of Sec20 is essential for yeast viability. Yeast strains lacking endogenous Sec20 were maintained using a WT Sec20 covering plasmid marked with Ura3 and a second plasmid with the His3-linked Sec20 allele (SC - HIS) indicated at left. When the Ura3 plasmid is lost on 5-fluoroorotic acid (5-FOA) selective plates (right), yeast harboring Sec20 lacking its N-terminal domain (ΔNTD) were inviable at all temperatures tested. Structure-based mutations in the Tip20•Sec20 interface (Tip20 V82D/L149D), which increased the equilibrium dissociation constant by ∼15-fold (Table S2), did not have an evident impact on viability.