FIGURE 5.
Shortened linker variants can be rescued by artificial linker sequences. A, influence of Hsp90 supplemented linker variants on viability of S. cerevisiae. The strains carrying the respective Hsp90 variant were plated on 5′-FOA-HIS agar and incubated for 4 days at 30 °C. B, the interaction of p23/Sba1 with the Hsp90 constructs was monitored by SPR at 25 °C in 40 mm Hepes, pH 7.5, 150 mm KCl, 5 mm MgCl2, 1 mm AMP-PNP. Binding of 1 μm of Hsp90 wt (—), Hsp90 Δ211–266SUP (… . .), or Hsp90 Δ211–272SUP (---) to immobilized p23/Sba1 was measured with injections of 65 μl. C, the ability of Hsp90 wt (●), Hsp90 Δ211–266SUP (○), or Hsp90 Δ211–272SUP (▾) to undergo N-terminal dimerization was investigated using the described FRET assay in 40 mm HEPES, 150 mm KCl, 5 mm MgCl2 at pH 7.5. The addition of ATPγS was used to start the assay.