New Generic Method for the Production of Protein-Based Inhibitors of Proteins Involved in Cancer Metastasis

摘要

Our objective was 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 used phage display and a display framework which was expected to bind preferentially to the active site pocket of target proteins. While we were able to find peptides in phage display libraries that bound to active stromelysin or to cadmium inactivated stromelysin and could find binders to a test target papain, in a constrained loop library we were unable to get the fully constrained scaffold protein to bind to its cognate target, subtilisin in the phage display system. Hence the overall objective was not obtained. As part of our efforts to characterize the binding epitope display framework, eglin c, we did develop a new method for using mutagenesis to study protein structure which we call patterned library analysis. We carried out a proof-of-principle experiment using the new method in which we were able to reproduce the values for alpha-helix propensity indicating that the method does indeed work. This method should have broad utility as a method for the assessment of hypotheses concerning the determinants of protein structure.