Year-end Sale % OFF 
Abstract
Saturation mutagenesis is a widely used directed evolution technique, in which a large number of protein variants, each having random amino acids in certain predetermined positions, are screened in order to discover high-fitness variants among them. Several metrics for determining the library size (the number of variants screened) have been suggested in the literature, but none of them incorporates the actual fitness of the variants discovered in the experiment. We present the results of an extensive simulation study, which is based on probabilistic models for protein fitness landscape, and which investigates how the result of a saturation mutagenesis experiment – the fitness of the best variant discovered – varies as a function of the library size. In particular, we study the loss of fitness in the experiment: the difference between the fitness of the best variant discovered, and the fitness of the best variant in variant space. Our results are that the existing criteria for determining the library size are conservative, so smaller libraries are often satisfactory. Reducing the library size can save labor, time, and expenses in the laboratory.
Highlights
Directed evolution has emerged as an indispensable vehicle for engineering biocatalysts, therapeutics, and other protein-based tools
In this work we provide another argument in favor of smaller libraries – one which is related to an experimental procedure called saturation mutagenesis
We wish to emphasize that it is not the intention of this work that protein engineering experimentalists should repeat themselves any of the computational or mathematical procedures described below; rather, we aim to derive general guidelines that are applicable to a wide range of saturation mutagenesis experiments, and that can be used by those practitioners
Summary
Directed evolution has emerged as an indispensable vehicle for engineering biocatalysts, therapeutics, and other protein-based tools. A key concept in evolutionary biology is that of fitness – the capacity of an organism to survive and reproduce. This meaning of the term originates from the phrase ‘‘survival of the fittest,’’ coined by Herbert Spencer and adopted by Charles Darwin. Though the very essence of directed evolution protocols depends on screening large numbers of mutants, recent works increasingly advocate relatively small, yet carefully designed libraries. Such targeted libraries enjoy a significantly higher fraction of functional mutants, compared to completely random libraries, and achieve remarkable experimental results. In this work we provide another argument in favor of smaller libraries – one which is related to an experimental procedure called saturation mutagenesis
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
