See the Lecture Summaries to read about the discussions that took place around each week’s readings.
| Week | Topics | Readings |
|---|---|---|
| 1 | Introduction |
There are no required readings for the first session, but below are some links to short introductory videos for those interested. And (still optional) a technical review paper for a more in-depth introduction. We recommend at least looking at the figures, as the full text may take a lot of time. Chemical & Engineering News. “The 2018 Nobel Prize in Chemistry: Directed evolution & phage display–Speaking of Chemistry.” October 4, 2018. YouTube. iGEM at Stony Brook. “Directed Evolution: Queens iGEM & Stony Brook iGEM Collaboration.” Sept. 30, 2018. YouTube. The Nobel Prize. “Frances H. Arnold: Nobel Lecture in Chemistry 2018.” Dec. 8, 2018. YouTube. Packer, M.S., and Liu, D.R. Methods for the directed evolution of proteins. Nature Reviews Genetics. 2015. 16, 379–394. |
| 2 | In vitro Gene Diversification. |
Wong, T.S., Tee, K.L., Hauer, B., and Schwaneberg, U. Sequence saturation mutagenesis (SeSaM): a novel method for directed evolution. Nucleic Acids Res. 2004. 32 (2): e26. Bittker, J.A., Le, B.V., Liu, J.M., and Liu, D.R. Directed evolution of protein enzymes using nonhomologous random recombination. PNAS. 2004. 101 (18), 7011–7016. |
| 3 | In vivo Gene Diversification. |
Wang, L., Coyt Jackson, W., Steinbach, P.A., and Tsien, R.Y. Evolution of new nonantibody proteins via iterative somatic hypermutation. PNAS. 2004. 101 (48), 16745-16749. DiCarlo, J.E., Conley, A.J., Penttila, M., Jantti, J., Wang, H.H., Church, G.M. Yeast Oligo-Mediated Genome Engineering (YOGE). ACS Synthetic Biology. 2013. 2(12), 741-749. |
| 4 | Identifying Desired Phenotypes – Selection Methods |
Kaltenbach, M., Stein, V., and Hollfelder, F. SNAP Dendrimers: Multivalent Protein Display on Dendrimer-Like DNA for Directed Evolution. ChemBioChem. 2011. 12, 2208-2216. Lin, H., Tao, H., and Cornish, V. W. Directed Evolution of a Glycosynthase via Chemical Complementation. J. Am. Chem. Soc. 2004. 126, 46, 15051-15059. |
| 5 | Identifying Desired Phenotypes – Screening Methods |
Kintses, B., Hein, C., Mohamed, M.F., Fischlechner, M., Courtois, F., Laine, C., and Hollfelder, F. Picoliter Cell Lysate Assays in Microfluidic Droplet Compartments for Directed Enzyme Evolution. Chemistry & Biology. 2012. 19(8), 1001-1009. Si, T., Li, B., Comi, T.J., Wu, Y., Hu, P., Wu, Y., Min, Y., Mitchell, D.A., Zhao, H., and Sweedler, J.V. Profiling of Microbial Colonies for High-Throughput Engineering of Multistep Enzymatic Reactions via Optically Guided Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry. J. Am. Chem. Soc. 2017. 139(36), 12466-12473. |
| 6 | Continuous Evolution |
Esvelt, K.M., Carlson, J.C., and Liu, D.R. A System For the continuous directed evolution of biomolecules. Nature. 2011. 472, 499-503. Berman, C.M., Papa, L.J., Hendel, S. J., Moore, C.L., Suen, P.H., Weickhardt, A.F., Doan, N., Kumar, C.M., Uil, T.G., Butty, V.L., Hoeben, R.C., and Shoulders, M.D. An Adaptable Platform for Directed Evolution in Human Cells. J. Am. Chem. Soc. 2018. 140(51), 18093-18103. |
| 7 | Addressing Climate Change |
Gomez-Fernandez, B. J., Garcia-Ruiz, E., Martin-Diaz, J., Gomez de Santos, P., Santos-Moriano, P., Plou, F. J., Ballesteros, A., Garcia, M., Rodriguez, M., Risso, V. A., Sanchez-Ruiz, J. M., Whitney, S. M., and Alcalde, M. Directed -in vitro- evolution of Precambrian and extant Rubiscos. Scientific Reports 2018. 8, 5532. Scheffen, M., Marchal, D. G., Beneyton, T., Schuller, S.K., Klose, M., Diehl, C., Lehmann, J., Pfister, P., Carrillo, M., He, H., Aslan, S., Cortina, N. S., Claus, P., Bollschweiler, D., Baret, J.-C., Schuller, J. M., Zarzycki, J., Bar-Even, A., and Erb, T.J. A new-to-nature carboxylation module to improve natural and synthetic CO2 fixation. Nature Catalysis 2021. 4, 105–115. |
| 8 | Engineering Waste Solutions | Bell, E., Smithson, R., Kilbride, S., Foster, J., Hardy, F., Ramachandran, S., Tedstone, A., Haigh, S., Garforth, A., Day, P., Levy, C., Shaver, M., and Green, A. Directed Evolution of an Efficient and Thermostable PET Depolymerase. ChemRxiv. 2021. |
| 9 | Developing Therapeutics |
Shusta, E.V., Holler, P.D., Kieke, M.C., Kranz, D.M., and Wittrup, D.K. Directed evolution of a stable scaffold for T-cell receptor engineering. Nature Biotechnology. 2000. 18, 754-759. Gray, J.P., Uddin, M.N., Chaudhari, R., Sutton, M.N., Yang, H., Rask, P., Locke, H., Engel, B.J., Batistatou, N., Wang, J., Grindel, B.J., Bhattacharya, P., Gammon, S.T., Zhang, S., Piwnica-Worms, D., Kritzer, J.A., Lu, Z., Bast, R.C., and Milward, S.W. Directed evolution of cyclic peptides for inhibition of autophagy. Chem. Sci. 2021. 12, 3526-3543. |
| 10 | Understanding Evolutionary Processes |
Nyerges, Á., Csörgő, B., Draskovits, G., Kintses, B., Szili, P., Ferenc, G., Révész, T., Ari, E., Nagy, I., Bálint, B. and Vásárhelyi, B.M. Directed evolution of multiple genomic loci allows the prediction of antibiotic resistance. Proceedings of the National Academy of Sciences, 2018. 115(25), E5726-E5735. Morgenthaler, A.B., Kinney, W.R., Ebmeier, C.C., Walsh, C.M., Snyder, D.J., Cooper, V.S., Old, W.M. and Copley, S.D. Mutations that improve efficiency of a weak-link enzyme are rare compared to adaptive mutations elsewhere in the genome. Elife, 2019. 8, e53535. |
| 11 | Next Generation Techniques in Directed Evolution: Evolution Beyond the Genetic Code and Natural Starting Points |
Reddington, S. C., Baldwin, A. J., Thompson, R., Brancale, A., Tippmann, E. M., and Jones, D. D. Directed evolution of GFP with non-natural amino acids identifies residues for augmenting and photoswitching fluorescence. Chem. Sci. 2015. 6, 1159–1166. Wang, M. S., and Hecht, M. H. A Completely De Novo ATPase from Combinatorial Protein Design. JACS. 2020. 142, 15230–15234. |
| 12 | Next Generation Techniques in Directed Evolution: Evolving Biosynthetic Pathways and Leveraging Machine Learning in Directed Evolution |
Shen, Y-P., Fong, L.S., Yan, Z-B., and Liu, J-Z. Combining directed evolution of pathway enzymes and dynamic pathway regulation using a quorum-sensing circuit to improve the production of 4-hydroxyphenylacetic acid in Escherichia coli. Biotechnology for Biofuels. 2019. 12, 94. Shen, M.W., Zhao, K.T., and Liu, D. R. Reconstruction of evolving gene variants and fitness from short sequencing reads. Nat. Chem. Biol. 2021. 17, 1188-1198. Optional Video: Machine Learning Assisted Dir. Evo. of Viral Vectors and Microbial Opsins for Minimally Invasive Neuroscience (30 min talk from Prof. Viviana Gradinaru at Caltech) |
| 13 | Final Presentations | No Readings |
