The ATPase cycle of Hsp90 drives a molecular 'clamp' via transient dimerization of the N-terminal domains

How the ATPase activity of Heat shock protein 90 (Hsp90) is coupled to clie nt protein activation remains obscure. Using truncation and missense mutant s of Hsp90, we analysed the structural implications of its ATPase cycle. C- terminal truncation mutants lacking inherent dimerization displayed reduced ATPase activity, but dimerized in the presence of 5'-adenyl-amido-diphosph ate (AMP-PNP), and AMP-PNP-promoted association of N-termini in intact Hsp9 0 dimers was demonstrated. Recruitment of p23/Sba1 to C-terminal truncation mutants also required AMPPNP-dependent dimerization. The temperature-sensi tive (ts) mutant T1O1I had normal ATP affinity but reduced ATPase activity and AMP-PNP-dependent N-terminal association, whereas the ts mutant T22I di splayed enhanced ATPase activity and AMPPNP-dependent N-terminal dimerizati on, indicating a close correlation between these properties. The locations of these residues suggest that the conformation of the 'lid' segment (resid ues 100-121) couples ATP binding to N-terminal association. Consistent with this, a mutation designed to favour 'lid' closure (A107N) substantially en hanced ATPase activity and N-terminal dimerization, These data show that Hs p90 has a molecular 'clamp' mechanism, similar to DNA gyrase and MutL, whos e opening and closing by transient N-terminal dimerization are directly cou pled to the ATPase cycle.