Engineering Environmentally-Stable Proteases to Specifically Neutralize Protein Toxins
Abstract
This project is intended to develop the tools and principles necessary to engineer subtilisin proteases which specifically target and deactivate biological warfare agent (BWA) toxins. We are engineering and evolving subtilisin proteases that specifically target and deactivate BoNT, SEB, ricin, and B. anthracis lethal factor (LF), representing four functionally distinct families of toxins. The centerpiece of our design effort is a phage-display selection method for creating tightly-regulated proteases of high specificity. In this system the protease, substrate sequence, and regulatory co-factor are co-evolved. The key accomplishments this past year were: 1) Design/evolution of a highly active enzyme that can cut P4 = I; 2) Design/evolution of an enzyme which can efficiently cut at P1 = Q; 3) First demonstration of the evolution of specificity for an ionic P4 amino acid ( P4 = E); 4) Engineering protease chain reactions that can reliably measure concentrations in the 0.1 to 10 pM range.
Document Details
- Document Type
- Technical Report
- Publication Date
- Oct 14, 2012
- Accession Number
- ADA584085
Entities
People
- Philip N. Bryan