Rapid Access to Antibiotic Biosynthesis Machinery Using Synthetic Biology

We will work with the Broad Institute and project leader Rob Nicol to deliver a new cell-free pipeline for moving from sequence to molecules. This will be pursued through three aims, to access three classes of antibiotics. • Aim 1: Express potential microbiome antibiotic gene clusters guided by genetic screen targets. We will mine microbiome sequences for potential antimicrobial gene clusters guided by targets from Drs. Hung and Balskus, which will then be expressed individually in our pipeline. • Aim 2. Rapid cell-free prototyping of stable cyclic peptides targeting gram negative enzymes. We will build large variant libraries (>108) of protegrin-1 analogues in droplets targeting LptD expressed in our pipeline to screen for function but also reduced human cell toxicity. • Aim 3. Rapid cell-free prototyping of lectins targeting microbial glycans. We will build large variant libraries of human lectin analogues targeting pathogen glycan structures.