High-throughput Droplet-based Microfluidics for Directed Evolution of Enzymes

Overview

Journal Electrophoresis

Specialty Chemistry

Date 2019 Aug 22

PMID 31433062

Citations 13

Authors

Flora W Y Chiu

Stavros Stavrakis

Affiliations

Soon will be listed here.

Abstract

Natural enzymes have evolved over millions of years to allow for their effective operation within specific environments. However, it is significant to note that despite their wide structural and chemical diversity, relatively few natural enzymes have been successfully applied to industrial processes. To address this limitation, directed evolution (DE) (a method that mimics the process of natural selection to evolve proteins toward a user-defined goal) coupled with droplet-based microfluidics allows the detailed analysis of millions of enzyme variants on ultra-short timescales, and thus the design of novel enzymes with bespoke properties. In this review, we aim at presenting the development of DE over the last years and highlighting the most important advancements in droplet-based microfluidics, made in this context towards the high-throughput demands of enzyme optimization. Specifically, an overview of the range of microfluidic unit operations available for the construction of DE platforms is provided, focusing on their suitability and benefits for cell-based assays, as in the case of directed evolution experimentations.

Citing Articles

Droplet Microfluidics for High-Throughput Screening and Directed Evolution of Biomolecules.

Vladisaljevic G Micromachines (Basel). 2024; 15(8).

PMID: 39203623 PMC: 11356158. DOI: 10.3390/mi15080971.

High-Throughput Absorbance-Activated Droplet Sorting for Engineering Aldehyde Dehydrogenases.

Jain A, Teshima M, Buryska T, Romeis D, Haslbeck M, Doring M Angew Chem Int Ed Engl. 2024; 63(49):e202409610.

PMID: 39087463 PMC: 11586695. DOI: 10.1002/anie.202409610.

Flow Cytometry: The Next Revolution.

Robinson J, Ostafe R, Narayana Iyengar S, Rajwa B, Fischer R Cells. 2023; 12(14).

PMID: 37508539 PMC: 10378642. DOI: 10.3390/cells12141875.

One cell at a time: droplet-based microbial cultivation, screening and sequencing.

Hu B, Xu P, Ma L, Chen D, Wang J, Dai X Mar Life Sci Technol. 2023; 3(2):169-188.

PMID: 37073344 PMC: 10077293. DOI: 10.1007/s42995-020-00082-8.

Biologically engineered microbes for bioremediation of electronic waste: Wayposts, challenges and future directions.

Han P, Teo W, Yew W Eng Biol. 2023; 6(1):23-34.

PMID: 36968558 PMC: 9995160. DOI: 10.1049/enb2.12020.

References

Song H, Ismagilov R . Millisecond kinetics on a microfluidic chip using nanoliters of reagents. J Am Chem Soc. 2003; 125(47):14613-9. PMC: 1769313. DOI: 10.1021/ja0354566. View

Abate A, Hung T, Mary P, Agresti J, Weitz D . High-throughput injection with microfluidics using picoinjectors. Proc Natl Acad Sci U S A. 2010; 107(45):19163-6. PMC: 2984161. DOI: 10.1073/pnas.1006888107. View

Peisajovich S, Tawfik D . Protein engineers turned evolutionists. Nat Methods. 2007; 4(12):991-4. DOI: 10.1038/nmeth1207-991. View

Pekin D, Skhiri Y, Baret J, Le Corre D, Mazutis L, Salem C . Quantitative and sensitive detection of rare mutations using droplet-based microfluidics. Lab Chip. 2011; 11(13):2156-66. DOI: 10.1039/c1lc20128j. View

Zhu Y, Fang Q . Analytical detection techniques for droplet microfluidics--a review. Anal Chim Acta. 2013; 787:24-35. DOI: 10.1016/j.aca.2013.04.064. View

Nge P, Rogers C, Woolley A . Advances in microfluidic materials, functions, integration, and applications. Chem Rev. 2013; 113(4):2550-83. PMC: 3624029. DOI: 10.1021/cr300337x. View

Guo M, Rotem A, Heyman J, Weitz D . Droplet microfluidics for high-throughput biological assays. Lab Chip. 2012; 12(12):2146-55. DOI: 10.1039/c2lc21147e. View

Courtois F, Olguin L, Whyte G, Theberge A, Huck W, Hollfelder F . Controlling the retention of small molecules in emulsion microdroplets for use in cell-based assays. Anal Chem. 2009; 81(8):3008-16. DOI: 10.1021/ac802658n. View

Najah M, Mayot E, Mahendra-Wijaya I, Griffiths A, Ladame S, Drevelle A . New glycosidase substrates for droplet-based microfluidic screening. Anal Chem. 2013; 85(20):9807-14. DOI: 10.1021/ac4022709. View

10.

Abate A, Poitzsch A, Hwang Y, Lee J, Czerwinska J, Weitz D . Impact of inlet channel geometry on microfluidic drop formation. Phys Rev E Stat Nonlin Soft Matter Phys. 2009; 80(2 Pt 2):026310. DOI: 10.1103/PhysRevE.80.026310. View

11.

Liau A, Karnik R, Majumdar A, Cate J . Mixing crowded biological solutions in milliseconds. Anal Chem. 2005; 77(23):7618-25. DOI: 10.1021/ac050827h. View

12.

Ostafe R, Prodanovic R, Commandeur U, Fischer R . Flow cytometry-based ultra-high-throughput screening assay for cellulase activity. Anal Biochem. 2012; 435(1):93-8. DOI: 10.1016/j.ab.2012.10.043. View

13.

Gupta R, Goldsmith M, Ashani Y, Simo Y, Mullokandov G, Bar H . Directed evolution of hydrolases for prevention of G-type nerve agent intoxication. Nat Chem Biol. 2011; 7(2):120-5. DOI: 10.1038/nchembio.510. View

14.

Kintses B, van Vliet L, Devenish S, Hollfelder F . Microfluidic droplets: new integrated workflows for biological experiments. Curr Opin Chem Biol. 2010; 14(5):548-55. DOI: 10.1016/j.cbpa.2010.08.013. View

15.

Gruner P, Riechers B, Semin B, Lim J, Johnston A, Short K . Controlling molecular transport in minimal emulsions. Nat Commun. 2016; 7:10392. PMC: 4735829. DOI: 10.1038/ncomms10392. View

16.

Colin P, Kintses B, Gielen F, Miton C, Fischer G, Mohamed M . Ultrahigh-throughput discovery of promiscuous enzymes by picodroplet functional metagenomics. Nat Commun. 2015; 6:10008. PMC: 4686663. DOI: 10.1038/ncomms10008. View

17.

Franke T, Abate A, Weitz D, Wixforth A . Surface acoustic wave (SAW) directed droplet flow in microfluidics for PDMS devices. Lab Chip. 2009; 9(18):2625-7. DOI: 10.1039/b906819h. View

18.

Romero P, Arnold F . Exploring protein fitness landscapes by directed evolution. Nat Rev Mol Cell Biol. 2009; 10(12):866-76. PMC: 2997618. DOI: 10.1038/nrm2805. View

19.

Fenneteau J, Chauvin D, Griffiths A, Nizak C, Cossy J . Synthesis of new hydrophilic rhodamine based enzymatic substrates compatible with droplet-based microfluidic assays. Chem Commun (Camb). 2017; 53(39):5437-5440. DOI: 10.1039/c7cc01506b. View

20.

Fischlechner M, Schaerli Y, Mohamed M, Patil S, Abell C, Hollfelder F . Evolution of enzyme catalysts caged in biomimetic gel-shell beads. Nat Chem. 2014; 6(9):791-6. DOI: 10.1038/nchem.1996. View