Protein Design, Protein Recognition and Molecular Chaperones
EMSL Project ID
2003
Abstract
Molecular chaperones such as heat shock proteins assist in protein folding by preventing misfolding and aggreagation, but also provide other vital cellular tasks such as import, translocation and degradation of proteins. A new protein BAG-1 binds to heat shock proteins and modulates the molecular chaperone activity of Hsp70. Bag proteins also bind to nuclear hormone receptors and may modulate the activity of these receptors by interaction with the heat shock molecular chaperones. In human tumor cell lines, levels of BAG-1L are elevated in hormone dependent cancers such as prostate, breast and lymphoid malignancies. It is in this area that BAG proteins may represent a novel target for future cancer therapies. BAG-1 may be a link between androgen receptor or estrogen receptor and heat shock chaperones. It has been suggested that BAG proteins and the cell death pathway could be new drug targets for cancer patients who are no longer responding to endocrine therapy. From earlier homology modeling of BAG-1/Hsc70 we predicted a binding crevice on Hsc70 as the site for BAG-1 interaction and confirmed this by NMR chemical shift experiments followed by mutagenesis. From these results a 'mini'domain of the Hsc70 ATPase domain has been produced by recombinant methods that contains the interaction sites in BAG recognition. This fragment is 60% helical. The plan is to determine the stucture of the 'mini' domain by NMR methods and to determine the structure of the 'mini'domain in complex with BAG-1. NMR experiments will be carried out with one partner unlabeled and the other partner labeled. The stategy is to use 15N-filtered and/or 13C,15N-filtered TROSY experiments. Our experience with the BAG-1 protein which is highly helical demonstrates that high field instsruments are required to resolve spectral ambiguities. We are requesting time for data collection on the 600MHz instrument and on the 750 MHz instrument.
Project Details
Project type
Capability Research
Start Date
2000-09-29
End Date
2001-03-02
Status
Closed
Released Data Link
Team
Principal Investigator