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Updated: Feb 6, 2026
Studies of Chaperone-Cochaperone Interactions using Homogenous Bead-Based Assay
Published on: July 21, 2021
Expression, Functional Characterization, and Preliminary Crystallization of the Cochaperone Prefoldin from the
Kento Morita1, Yohei Y Yamamoto2, Ayaka Hori3
1Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, Naka, Koganei, Tokyo 184-8588, Japan. kento.morita@yohda.net.
Abstract:
Prefoldin is a hexameric molecular chaperone found in the cytosol of archaea and eukaryotes. Its hexameric complex is built from two related classes of subunits, and has the appearance of a jellyfish: Its body consists of a double β-barrel assembly with six long tentacle-like coiled coils protruding from it. Using the tentacles, prefoldin captures an unfolded protein substrate and transfers it to a group II chaperonin. Based on structural information from archaeal prefoldins, mechanisms of substrate recognition and prefoldin-chaperonin cooperation have been investigated. In contrast, the structure and mechanisms of eukaryotic prefoldins remain unknown. In this study, we succeeded in obtaining recombinant prefoldin from a thermophilic fungus, Chaetomium thermophilum (CtPFD). The recombinant CtPFD could not protect citrate synthase from thermal aggregation. However, CtPFD formed a complex with actin from chicken muscle and tubulin from porcine brain, suggesting substrate specificity. We succeeded in observing the complex formation of CtPFD and the group II chaperonin of C. thermophilum (CtCCT) by atomic force microscopy and electron microscopy. These interaction kinetics were analyzed by surface plasmon resonance using Biacore. Finally, we have shown the transfer of actin from CtPFD to CtCCT. The study of the folding pathway formed by CtPFD and CtCCT should provide important information on mechanisms of the eukaryotic prefoldin⁻chaperonin system.
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