Ultrahigh-throughput-directed enzyme evolution by absorbance-activated droplet sorting (AADS)
Abstract
Directed enzyme evolution is a powerful approach for discovering new catalysts with applications in green chemistry and elsewhere. However, "hits" in sequence space are rare: If too few members of a library are examined, the chances of success of a campaign are limited. Ultrahigh-throughput screening in emulsion droplets has dramatically increased the odds but requires a fluorescent reaction product that triggers selection of hits. We now introduce an absorbance-based microfluidic droplet sorter that broadens the scope of assays to those producing UV/Vis-active chromophores and demonstrate its usefulness by evolving a dehydrogenase based on the screening of half a million library members. Making ultrahigh-throughput screening possible for previously inaccessible reactions enables much wider use of microfluidic droplet sorters for laboratory evolution.
- Publication:
-
Proceedings of the National Academy of Science
- Pub Date:
- November 2016
- DOI:
- 10.1073/pnas.1606927113
- Bibcode:
- 2016PNAS..113E7383G
- Keywords:
-
- protein engineering;
- directed evolution;
- microfluidics;
- ultrahigh-throughput;
- emulsion droplets