A New Generic Method for the Production of Protein-Based Inhibitors of Proteins Involved in Cancer Metastasis
Abstract
The objective is to develop a method to make protein-based inhibitors against protein targets and as the test case to a proteinase involved in metastasis, stromelysin. The approach uses phage display and a display framework which should bind preferentially to the active site pocket of target proteins. We have now turned to construction technology which can be applied to any portion of the framework independent of restriction site distribution. A modified vector has been constructed and sequenced to support this approach. We have shown that peptides that bind to active stromelysin also bind to cadmium inactivated stromelysin which will not cleave weak binders allowing us to detect them and work to improve their affinity. In an effort to characterize the binding epitope display framework, eglin c, we have also developed a new method for using mutagenesis to study protein structure which we call patterned library analysis. We have worked out a number of the technical details of employing this approach. In the process we have also discovered that eglin is reversibly denatured in low levels of SDS. This opens up the possibility of collecting thermodynamic parameters with an activity based assay that uses 100-fold less material than traditional biophysical techniques.
Document Details
- Document Type
- Technical Report
- Publication Date
- Aug 01, 1998
- Accession Number
- ADA366569
Entities
People
- Marshall H. Edgell
Organizations
- University of North Carolina at Chapel Hill