UNIVERSITY OF MASSACHUSETTS
Hsp90 is a unique chaperone that is essential in eukaryotes and that helps to produce and maintain the active state of a select set of biologically and medically important substrates/clients including many signal transduction proteins. Through these clients, Hsp90 is involved in biological processes including aging, signal transduction and evolution. Hsp90 function requires ATP hydrolysis and the dynamic binding and release of clients and numerous co-chaperones. This type of dynamic macromolecular assembly process underlies many critical biological processes including DNA replication and the initiation of transcription. Understanding the conformational dynamics of Hsp90 will provide insights into other dynamic macromolecular complexes and determine the role of chaperones in signal transduction. Many different conformational cycles of Hsp90 are possible based on the biochemical properties of Hsp90. We are elucidating the biologically relevant Hsp90 conformations in vivo. We use protein engineering strategies to thermodynamically stabilize Hsp90 in distinct conformations in order to determine their biochemical properties and their function in vivo. The results of these experiments will delineate the Hsp90 conformations that activate clients in vivo and determine the biologically relevant Hsp90 chaperone cycle. The purpose of this supplement is to complement our analyses of the v-src kinase with quantitative studies of a panel of endogenous yeast kinases. Examining the role of Hsp90 in the maturation of multiple kinases will provide insight into both general and variable mechanistic aspects of the chaperone system. If our modulations of Hsp90 conformation impact all kinases in a similar fashion that would indicate that Hsp90 matures all kinases through a similar mechanism and justify our initial approach of using v-src maturation as a paradigm for all kinases. On the other hand, Hsp90 contains multiple hydrophobic surface patches capable of binding to kinases in different orientations and leading to kinase maturation through different chaperone/kinase complexes. In this case our modulations of Hsp90 conformation would differentially impact the maturation of kinases depending on their interaction mode with Hsp90.