Directed Evolution. The Legacy of a Nobel Prize


This article is part of an anniversary issue of Journal of Molecular Evolution, commenting on a paper published on 1999 by the Nobel laureate Frances Arnold and her colleague Kentaro Miyazaki. The paper by Miyazaki and Arnold presented saturation mutagenesis as an alternative method to random mutagenesis for obtaining enzymes with increasing stability. Both techniques were conceived to accomplish directed evolution, an approach honoured by the Nobel Prize of Chemistry 2018. Here, I am commenting on the pros and cons of random and saturation mutagenesis, while also discussing important results from directed evolution. I conclude that molecular evolution is finding new applications in science and it is definitely an integral part of the genomic era’s revolution.

This is a preview of subscription content, access via your institution.


  1. Arnold FH (2019) Innovation by evolution: bringing new chemistry to life (Nobel lecture). Angew Chem Int Ed 58:14420–14426

    Article  CAS  Google Scholar 

  2. Bloom JD, Arnold FH (2009) In the light of directed evolution: pathways of adaptive protein evolution. Proc Natl Acad Sci USA 106(Suppl):9995–10000

    Article  Google Scholar 

  3. Christaki E, Marcou M, Tofarides A (2020) Antimicrobial resistance in bacteria: mechanisms, evolution, and persistence. J Mol Evol 88:26–40

    Article  CAS  Google Scholar 

  4. Coelho PS, Brustad EM, Kannan A, Arnold FH (2013) Olefin cyclopropanation via carbene transfer catalyzed by engineered cytochrome P450 enzymes. Science 339:307–310.

    Article  PubMed  CAS  Google Scholar 

  5. Colautti RI, Alexander JM, Dlugosch KM et al (2017) Invasions and extinctions through the looking glass of evolutionary ecology. Philos Trans R Soc B 372:20160031

    Article  Google Scholar 

  6. Davis BH, Poon AFY, Whitlock MC (2009) Compensatory mutations are repeatable and clustered within proteins. Proc R Soc B 276:1823–1827.

    Article  PubMed  Google Scholar 

  7. Gupta K, Varadarajan R (2018) Insights into protein structure, stability and function from saturation mutagenesis. Curr Opin Struct Biol 50:117–125

    Article  CAS  Google Scholar 

  8. Kan SBJ, Lewis RD, Chen K, Arnold FH (2016) Directed evolution of cytochrome c for carbon-silicon bond formation: bringing silicon to life. Science 354:1048–1051.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  9. Kimura M (1968) Evolutionary rate at the molecular level. Nature 217:624–626.

    Article  PubMed  CAS  Google Scholar 

  10. Lenski RE (2017) Experimental evolution and the dynamics of adaptation and genome evolution in microbial populations. ISME J 11:2181–2194

    Article  CAS  Google Scholar 

  11. Lenski RE, Rose MR, Simpson SC, Tadler SC (1991) Long-term experimental evolution in Escherichia coli. I. Adaptation and divergence during 2000 generations. Am Nat 138:1315–1341.

    Article  Google Scholar 

  12. Liberles DA, Chang B, Geiler-Samerotte K et al (2020) Emerging frontiers in the study of molecular evolution. J Mol Evol 88:211–226

    Article  CAS  Google Scholar 

  13. Maynard Smith J (1970) Natural selection and the concept of a protein space. Nature 225:563–564.

    Article  Google Scholar 

  14. McIntosh JA, Coelho PS, Farwell CC et al (2013) Enantioselective intramolecular C-H amination catalyzed by engineered cytochrome P450 enzymes in vitro and in vivo. Angew Chem Int Ed 52:9309–9312.

    Article  CAS  Google Scholar 

  15. Miyazaki K, Arnold FH (1999) Exploring nonnatural evolutionary pathways by saturation mutagenesis: rapid improvement of protein function. J Mol Evol 49:716–720.

    Article  PubMed  CAS  Google Scholar 

  16. Omura T (2013) Contribution of cytochrome P450 to the diversification of eukaryotic organisms. Biotechnol Appl Biochem 60:4–8

    Article  CAS  Google Scholar 

  17. Renata H, Wang ZJ, Arnold FH (2015) Expanding the enzyme universe: accessing non-natural reactions by mechanism-guided directed evolution. Angew Chem Int Ed 54:3351–3367

    Article  CAS  Google Scholar 

  18. Sayous V, Lubrano P, Li Y, Acevedo-Rocha CG (2020) Unbiased libraries in protein directed evolution. Biochim Biophys Acta 1868:140321

    Article  CAS  Google Scholar 

  19. Stearns SC (2020) Frontiers in molecular evolutionary medicine. J Mol Evol 88:3–11

    Article  CAS  Google Scholar 

  20. Wang ZJ, Renata H, Peck NE et al (2014) Improved cyclopropanation activity of histidine-ligated cytochromeP450 enables the enantioselective formal synthesis of levomilnacipran. Angew Chem Int Ed 53:6810–6813.

    Article  CAS  Google Scholar 

  21. Zahir N, Sun R, Gallahan D et al (2020) Characterizing the ecological and evolutionary dynamics of cancer. Nat Genet 52:759–767.

    Article  PubMed  CAS  Google Scholar 

  22. Zeymer C, Hilvert D (2018) Directed evolution of protein catalysts. Annu Rev Biochem 87:131–157

    Article  CAS  Google Scholar 

Download references


Not applicable.

Author information



Corresponding author

Correspondence to Konstantinos Voskarides.

Ethics declarations

Conflicts of interest

Not any.

Additional information

Handling editor: Aaron Goldman.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Voskarides, K. Directed Evolution. The Legacy of a Nobel Prize. J Mol Evol 89, 189–191 (2021).

Download citation


  • Natural selection
  • Mutation
  • Protein engineering
  • Enzymes
  • Medicine
  • Fitness
  • Neutral evolution