Structure of a Helical Signaling Domain from p130Cas
EMSL Project ID
2423
Abstract
The cellular responses to integrin signals are coordinated through specific protein-protein interactions. In this project we focus on an adaptor protein called Cas that becomes phosphorylated upon integrin/ligand binding, forms complexes with a number of signaling proteins and switches on and off downstream signals. The tyrosine-phosphorylation level of Cas correlates well with the transforming phenotype of cells and is thought to play an essential role in the process of cell transformation. Recently it has been recognized that Cas is identical to the BCAR-1 gene that causes resistance to anti-estrogen drugs (tamoxifen) in breast cancer cells. Adaptor proteins such as Cas interact with a number of different molecules. The interacting surfaces between these pairs of proteins are potential targets for drug design, but structural details of the partner molecules must be known. Cas contains a ‘serine-rich’ region that is serine-phosphorylated following integrin-mediatad cell adhesion. Serine-phosphorylation is important for interactions with 14-3-3 protein that plays a chaperone role to modulate signal transduction pathways. The Cas/14-3-3 complex may be involved in activation of the Erk pathway. The 3D structure of 14-3-3 is known and the goal of this project is to determine the structure of Cas serine-rich domain that binds to it. Secondary structure prediction and circular dichroism indicate that the domain is highly helical. Recombinant constructs are in hand to prepare soluble protein for NMR analyses. 1H-15N and 1H-13C HSQC spectra reveal that amide and aliphatic resonances are clustered, so the experiments will benefit from data acquisition at high fields to relieve the spectral overlap and improve the resolution. The plan is to record a set of 4D NOESY experiments using labeled protein to complete the assignments and to obtain unambiguous distance restraints for structure calculation.
Project Details
Project type
Capability Research
Start Date
2002-04-03
End Date
2002-08-30
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members
Related Publications
F Nasertorabi, Briknarova K, ML Havert, E Eggleston, DW Hoyt, C Li, AJ Olson, K Vuori, and KR Ely. 2005. "The Serine-rich Domain from Crk-associated Substrate (p130Cas) is a Four-helix Bundle." Journal of Biological Chemistry 280(23):21908-21914.