Prions are unconventional, highly infectious agents, which are composed entirely of a protein that adopts an abnormal conformation. In mammals, prion-mediated infections are responsible for several devastating and invariably fatal neurodegenerative diseases, collectively known as transmissible spongiform encephalopathies. A hallmark of prion diseases is the presence of amyloids, which are also associated with the pathology of non-prion diseases, ranging from Alzheimer's and Huntington's disease to systemic amyloidosis. The broad and long-term research objective is to uncover the functional role of molecular chaperones in prion replication. Yeast provides an excellent paradigm to investigate the mechanism of prion replication. [PSI+] is a yeast prion that increases translational read-through of nonsense codons. Like mammalian prions, [PSI+] consist entirely of protein and is formed by self-replicating amyloid conformers of the evolutionary conserved translation termination factor Sup35p (eRF3). The inheritance and maintenance of [PSI+] are governed by Hsp104, a 600-kDa, ring-forming ATP-dependent, protein-remodeling machine, which cooperates with the Hsp70 chaperone system in prion replication and protein disaggregation. The objective of this research is to provide a detailed mechanistic understanding of the prion-remodeling and protein disaggregating activities of Hsp104 and its bacterial homolog ClpB.
Three specific aims are proposed: 1) to determine the 3D structure of an Hsp104-substrate complex, 2) to investigate the synergistic interaction between Hsp104 and Hsp70/Hsp40, and 3) to elucidate the mechanism of protein disaggregation and prion replication by the Hsp104 bi-chaperone system. To address our research questions, we will use a multi-facet approach consisting of hybrid structural biology methods, proteomic and chemical biology techniques, and yeast genetics. The combination of these methods provides a powerful approach to yield new mechanistic insight into the structure-function relationship of this remarkable family of ATP-dependent molecular machines in order that this information might be exploited to engineer new nano-machines with novel biological activities with potential applications in biotechnology and nano-medicine.

Public Health Relevance

Prions are highly infectious, proteinaceous agents responsible for several devastating and invariably fatal neurodegenerative diseases. While molecular chaperones might be expected to provide the first line of defense against prion-mediated infections and other protein conformational disorders, their function (or lack thereof) in protein misfolding diseases is poorly understood. Here, we propose to investigate the structure, function, and mechanism of the ring-forming Hsp104 molecular chaperone, an ATP-dependent prion-remodeling machine and essential factor of the stress response.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM104980-06
Application #
8670000
Study Section
Special Emphasis Panel (ZRG1-BCMB-R (02))
Program Officer
Wehrle, Janna P
Project Start
2013-07-01
Project End
2017-03-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
6
Fiscal Year
2014
Total Cost
$316,913
Indirect Cost
$114,413
Name
Baylor College of Medicine
Department
Biochemistry
Type
Schools of Medicine
DUNS #
051113330
City
Houston
State
TX
Country
United States
Zip Code
77030
Lee, Jungsoon; Kim, Ji-Hyun; Biter, Amadeo B et al. (2013) Heat shock protein (Hsp) 70 is an activator of the Hsp104 motor. Proc Natl Acad Sci U S A 110:8513-8