title: Enforced N-domain proximity stimulates Hsp90 ATPase activity and is compatible with function in vivo.
abstract: Hsp90 populates distinct open and closed conformations mediated by transient N-terminal dimerization. To investigate the mechanistic role of these large conformational changes, we designed Hsp90 with an N-terminal coiled-coil to clamp the termini together and enforce N-domain proximity. Biophysical analyses demonstrate that the coiled-coil effectively maintains N-domain proximity in the absence of ATP, a condition that favors the open state of Hsp90. Enforcing N-domain proximity results in increased ATPase activity, indicating that N-terminal dimerization is a rate-limiting step that is sped-up with the coiled-coil due to increased effective N-domain concentration. The relative difference in ATPase activity between coil-Hsp90 and wt was reduced in the presence of both an ATPase activating (Aha1) and an inhibiting (Sba1) co-chaperone. As both of these co-chaperones bind preferentially to N-terminally dimerized Hsp90, the buffering effect of these co-chaperones demonstrates the biochemical relevance of Hsp90 conformational properties in addition to N-terminal dimerization. Enforcing N-domain proximity is compatible with viability in yeast, underlining the mechanistic relevance of Hsp90 conformational changes that are less dramatic than the transition between fully open and closed.
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA.
authors.forename: Lester
authors.initials: L
authors.lastname: Pullen
authors.forename: Daniel N
authors.initials: DN
authors.lastname: Bolon
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D055503:Protein Multimerization / Q000502:physiology*; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary
chemical_list: D018841:HSP90 Heat-Shock Proteins; D000251:Adenosine Triphosphatases