title: Intrinsic inhibition of the Hsp90 ATPase activity.
abstract: The molecular chaperone Hsp90 is required for the folding and activation of a large number of substrate proteins. These are involved in essential cellular processes ranging from signal transduction to viral replication. For the activation of its substrates, Hsp90 binds and hydrolyzes ATP, which is the key driving force for conformational conversions within the dimeric chaperone. Dimerization of Hsp90 is mediated by a C-terminal dimerization site. In addition, there is a transient ATP-induced dimerization of the two N-terminal ATP-binding domains. The resulting ring-like structure is thought to be the ATPase-active conformation. Hsp90 is a slow ATPase with a turnover number of 1 ATP/min for the yeast protein. A key question for understanding the molecular mechanism of Hsp90 is how ATP hydrolysis is regulated and linked to conformational changes. In this study, we analyzed the activation process structurally and biochemically with a view to identify the conformational limitations of the ATPase reaction cycle. We showed that the first 24 amino acids stabilize the N-terminal domain in a rigid state. Their removal confers flexibility specifically to the region between amino acids 98 and 120. Most surprisingly, the deletion of this structure results in the complete loss of ATPase activity and in increased N-terminal dimerization. Complementation assays using heterodimeric Hsp90 show that this rigid lid acts as an intrinsic kinetic inhibitor of the Hsp90 ATPase cycle preventing N-terminal dimerization in the ground state. On the other hand, this structure acts, in concert with the 24 N-terminal amino acids of the other N-terminal domain, to form an activated ATPase and thus regulates the turnover number of Hsp90.
authors.affiliation: Institut für Organische Chemie und Biochemie, Technische Universität München, Lichtenbergstrasse 4, 85747 Garching, Germany.
authors.forename: Klaus
authors.initials: K
authors.lastname: Richter
authors.forename: Sandra
authors.initials: S
authors.lastname: Moser
authors.forename: Franz
authors.initials: F
authors.lastname: Hagn
authors.forename: Rainer
authors.initials: R
authors.lastname: Friedrich
authors.forename: Otmar
authors.initials: O
authors.lastname: Hainzl
authors.forename: Markus
authors.initials: M
authors.lastname: Heller
authors.forename: Sandra
authors.initials: S
authors.lastname: Schlee
authors.forename: Horst
authors.initials: H
authors.lastname: Kessler
authors.forename: Jochen
authors.initials: J
authors.lastname: Reinstein
authors.forename: Johannes
authors.initials: J
authors.lastname: Buchner
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D000595:Amino Acid Sequence; D002942:Circular Dichroism; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004058:Diffusion; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D017353:Gene Deletion; D018841:HSP90 Heat-Shock Proteins / Q000037:antagonists & inhibitors*; D006868:Hydrolysis; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D015398:Signal Transduction; D013379:Substrate Specificity; D013997:Time Factors; D014508:Urea / Q000494:pharmacology
chemical_list: D004791:Enzyme Inhibitors; D018841:HSP90 Heat-Shock Proteins; D011506:Proteins; D000255:Adenosine Triphosphate; D014508:Urea; D000251:Adenosine Triphosphatases