Abstract
While the molecular details by which Hsp90 interacts with Sgt1 and Rar1 were previously described the exact stoichiometric complex that is formed remains elusive. Several possibilities remain that include two asymmetric complexes, Sgt1(2)-Hsp90(2)-Rar1(2) (two molecules of Sgt1 and Rar1 and one Hsp90 dimer) or Sgt1(2)-Hsp90(2)-Rar1(1) (with a single Rar1 molecule) and an asymmetric complex (Sgt1(1)-Hsp90(2)-Rar1(1)). The Hsp90-mediated activation of NLR receptors (Nucleotide-binding domain and Leucine-rich Repeat) in the innate immunity of both plants and animals is dependent on the co-chaperone Sgt1 and in plants on Rar1, a cysteine- and histidine-rich domain (CHORD)-containing protein. The exact stoichiometry of such a complex may have a direct impact on NLR protein oligomerization and thus ultimately on the mechanism by which NLRs are activated. CD spectroscopy was successfully used to determine the stoichiometry of a ternary protein complex among Hsp90, Sgt1, and Rar1 in the presence of excess ADP. The results indicated that a symmetric Sgt1(2)-Hsp90(2)-Rar1(1) complex was formed that could allow two NLR molecules to simultaneously bind. The stoichiometry of this complex has implications on, and might promote, the dimerization of NLR proteins following their activation.