Bacteriophage Engineering: L-Protein Optimization 1. Hypothesis: Engineering Lysis Protein Stability Our core hypothesis is that the thermodynamic stability and lytic efficiency of the MS2 L-protein can be enhanced through two strategic pathways:
Structural Reinforcement: Introducing targeted mutations that promote independent folding or stabilize the 7-helix bundle, reducing dependence on the host chaperone DnaJ. Generative Optimization: Utilizing evolutionary conservation data and generative protein design to create variants with improved membrane-insertion kinetics and host compatibility, thereby minimizing host-mediated resistance. 2. Specific Aims and Validation Pipeline Aim 1: Mutation Design via Conservation and Predictive Modeling We will perform Clustal Omega alignments of homologous lysis proteins to identify conserved residues (specifically the “HEDYPCRRQQRSST” island). This is followed by: