Fig. 1.

Interaction of PalF with the PalH C-terminal cytoplasmic tail. (A) Predicted PalH seven-pass topology and C-terminal cytoplasmic tail. (B) Overlapping fragments of the PalH cytoplasmic tail obtained in two-hybrid screening by using PalF bait. Positive interactions were revealed by growth of S. cerevisiae TAT7 on histidine-free medium containing 5 mM 3-aminotriazole (His⁺) and β-galactosidase lift filter assays (β-gal). (C) In vitro binding of PalF to the PalH cytoplasmic tail. GST-PalF or GST-PalA was immobilized on glutathione-Sepharose beads and incubated with in-vitro-synthesized [³⁵S]PalH (349-760). Pulled down proteins were electrophoresed on 10% SDS/polyacrylamide and detected by autoradiography (Upper) and Coomassie staining (Lower). Input: purified GST fusion proteins (Coomassie) or [³⁵S]PalH (349-760) (autoradiography) used for binding experiments (20% of the total reaction mixture).