Helena Rubinstein Professor, Department of Chemistry, Columbia University
In vitro directed evolution allows biomolecules with new and useful properties to be engineered—mimicking natural evolution on an experimentally accessible time scale by creating large libraries of DNA mutants using PCR and then carrying out a high-throughput assay for variants with improved function. To provide a breakthrough in the complexity of libraries that can be readily searched experimentally for systems engineering, my laboratory is engineering S. cerevisiae so that both the mutagenesis and selection steps of directed evolution can be carried out entirely in vivo, under conditions of sexual reproduction. I will describe chemical complementation, which provides a selection for chemistry beyond that natural to the cell using themes and variations on the yeast two-hybrid assay. Then I will present a heritable recombination system, for simultaneous mutagenesis and selection in vivo under conditions of sexual reproduction. Finally, the generality of this approach is explored for protein and systems engineering.