Structural Studies of the pRB Tumor Suppressor Complexed with Human Papillomavirus E7.
Abstract
Since viral oncoproteins are expected to compete with and imitate interactions that pRB has with cyclin D1, understanding high affinity pRB-viral oncoprotein complexes will provide tremendous insight into the specific interactions required for the development of small compounds that can destabilize pRB-cyclin D1 complexes in cyclin D1 mediated breast cancer. A structure-based approach is used to understand the molecular mechanisms that are necessary for high affinity pRB-viral oncoprotein interaction. The recently determined pRB-E7 peptide crystal structure31 has significant shortcomings because the crystallized E7 nine amino acid peptide binds to pRB with a twenty-fold weaker affinity compared to full-length HPV16 E732. Therefore, the goal of this project is to determine the three dimensional structure of pRB hound to larger regions of HPV E7, and Adenovirus 5 E1A. This study demonstrated that bacterially coexpressed pRB(376-792) and viral oncoproteins formed complexes. These purified complexes probably have resisted crystallization because pRB(376-792) included a flexible linker region between two structured domains of pRB. Partial proteolysis of pRB-viral oncoprotein complexes yielded three protein fragments that are consistent with the molecular weights of pRB structured domains and larger viral oncoprotein regions (which contain high affinity pRB binding domains). Therefore, we expect that limited proteolysis will result in crystallizable pRB-viral oncoprotein complexes.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jun 01, 1998
- Accession Number
- ADA354361
Entities
People
- Adrienne M. Clements
Organizations
- University of Pennsylvania