Content area
Full Text
REVIEWS
S T U DY D E S I G N S
Methods for the directed evolution of proteins
Michael S.Packer and David R.Liu
Abstract | Directed evolution has proved to be an effective strategy for improving or altering the activity of biomolecules for industrial, research and therapeutic applications. The evolution of proteins in the laboratory requires methods for generating genetic diversity and for identifying protein variants with desired properties. This Review describes some of the tools used to diversify genes, as well as informative examples of screening and selection methods that identify or isolate evolved proteins. We highlight recent cases in which directed evolution generated enzymatic activities and substrate specificities not known to exist in nature.
Natural selection
A process by which individuals with the highest reproductive fitness pass on their genetic material to their offspring, thus maintaining and enriching heritable traits that are adaptive to the natural environment.
Artificial selection
(Also known as selective breeding). A process by which human intervention in the reproductive cycle imposes a selection pressure for phenotypic traits desired by the breeder.
Libraries
Diverse populations of DNA fragments that are subject to downstream screening and selection.
Over many generations, iterated mutation and natural selection during biological evolution provide solutions for challenges that organisms face in the natural world. However, the traits that result from natural selection only occasionally overlap with features of organisms and biomolecules that are sought by humans. To guide evolution to access useful phenotypes more frequently, humans for centuries have used artificial selection, beginning with the selective breeding of crops1 and domestication of animals2. More recently, directed evolution in the laboratory has proved to be a highly effective and broadly applicable framework for optimizing or altering the activities of individual genes and gene products, which are the fundamental units of biology.
Genetic diversity fuels both natural and laboratory evolution. The occurrence rate of spontaneous mutations is generally insufficient to access desired gene variants on a time scale that is practical for laboratory evolution. A number of genetic diversification techniques are therefore used to generate libraries of gene variants that accelerate the exploration of a genes sequence space. Methods to identify and isolate library members with desired properties are a second crucial component of laboratory evolution. During...