“NAD‐display”: Ultrahigh‐Throughput in Vitro Screening of NAD(H) Dehydrogenases Using Bead Display and Flow Cytometry
NAD(H)‐utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell‐free, ultrahigh‐throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per s...
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| Vydáno v: | Angewandte Chemie International Edition Ročník 60; číslo 16; s. 9015 - 9021 |
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Wiley Subscription Services, Inc
12.04.2021
John Wiley and Sons Inc |
| Vydání: | International ed. in English |
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| ISSN: | 1433-7851, 1521-3773, 1521-3773 |
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| Abstract | NAD(H)‐utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell‐free, ultrahigh‐throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per sample) carrying both variant DNA and an immobilised analogue of NAD+ were compartmentalised in water‐in‐oil emulsion droplets, together with cell‐free expression mixture and enzyme substrate, resulting in the recording of the phenotype on each bead. The beads’ phenotype could be read out and sorted for on a flow cytometer by using a highly sensitive fluorescent protein‐based sensor of the NAD+:NADH ratio. Integration of this “NAD‐display” approach with our previously described Split & Mix (SpliMLiB) method for generating large site‐saturation libraries allowed straightforward screening of fully balanced site saturation libraries of formate dehydrogenase, with diversities of 2×104. Based on modular design principles of synthetic biology NAD‐display offers access to sophisticated in vitro selections, avoiding complex technology platforms.
A detection system was devised to screen dehydrogenase enzymes (DH) at ultrahigh‐throughput in in vitro droplet compartments. An NAD(H) analogue and a DH DNA library were co‐immobilised on beads. Water‐in‐oil emulsion droplets containing cell‐free expression mix and substrate allow compartmentalised protein production and catalysis. A fluorescent sensor of NAD(H) redox state was loaded onto beads, allowing bulk sorting of 2×104 DH variants in a day by flow cytometry. |
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| AbstractList | NAD(H)-utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell-free, ultrahigh-throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per sample) carrying both variant DNA and an immobilised analogue of NAD+ were compartmentalised in water-in-oil emulsion droplets, together with cell-free expression mixture and enzyme substrate, resulting in the recording of the phenotype on each bead. The beads' phenotype could be read out and sorted for on a flow cytometer by using a highly sensitive fluorescent protein-based sensor of the NAD+ :NADH ratio. Integration of this "NAD-display" approach with our previously described Split & Mix (SpliMLiB) method for generating large site-saturation libraries allowed straightforward screening of fully balanced site saturation libraries of formate dehydrogenase, with diversities of 2×104 . Based on modular design principles of synthetic biology NAD-display offers access to sophisticated in vitro selections, avoiding complex technology platforms.NAD(H)-utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell-free, ultrahigh-throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per sample) carrying both variant DNA and an immobilised analogue of NAD+ were compartmentalised in water-in-oil emulsion droplets, together with cell-free expression mixture and enzyme substrate, resulting in the recording of the phenotype on each bead. The beads' phenotype could be read out and sorted for on a flow cytometer by using a highly sensitive fluorescent protein-based sensor of the NAD+ :NADH ratio. Integration of this "NAD-display" approach with our previously described Split & Mix (SpliMLiB) method for generating large site-saturation libraries allowed straightforward screening of fully balanced site saturation libraries of formate dehydrogenase, with diversities of 2×104 . Based on modular design principles of synthetic biology NAD-display offers access to sophisticated in vitro selections, avoiding complex technology platforms. NAD(H)‐utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell‐free, ultrahigh‐throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per sample) carrying both variant DNA and an immobilised analogue of NAD + were compartmentalised in water‐in‐oil emulsion droplets, together with cell‐free expression mixture and enzyme substrate, resulting in the recording of the phenotype on each bead. The beads’ phenotype could be read out and sorted for on a flow cytometer by using a highly sensitive fluorescent protein‐based sensor of the NAD + :NADH ratio. Integration of this “NAD‐display” approach with our previously described Split & Mix (SpliMLiB) method for generating large site‐saturation libraries allowed straightforward screening of fully balanced site saturation libraries of formate dehydrogenase, with diversities of 2×10 4 . Based on modular design principles of synthetic biology NAD‐display offers access to sophisticated in vitro selections, avoiding complex technology platforms. NAD(H)‐utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell‐free, ultrahigh‐throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per sample) carrying both variant DNA and an immobilised analogue of NAD+ were compartmentalised in water‐in‐oil emulsion droplets, together with cell‐free expression mixture and enzyme substrate, resulting in the recording of the phenotype on each bead. The beads’ phenotype could be read out and sorted for on a flow cytometer by using a highly sensitive fluorescent protein‐based sensor of the NAD+:NADH ratio. Integration of this “NAD‐display” approach with our previously described Split & Mix (SpliMLiB) method for generating large site‐saturation libraries allowed straightforward screening of fully balanced site saturation libraries of formate dehydrogenase, with diversities of 2×104. Based on modular design principles of synthetic biology NAD‐display offers access to sophisticated in vitro selections, avoiding complex technology platforms. A detection system was devised to screen dehydrogenase enzymes (DH) at ultrahigh‐throughput in in vitro droplet compartments. An NAD(H) analogue and a DH DNA library were co‐immobilised on beads. Water‐in‐oil emulsion droplets containing cell‐free expression mix and substrate allow compartmentalised protein production and catalysis. A fluorescent sensor of NAD(H) redox state was loaded onto beads, allowing bulk sorting of 2×104 DH variants in a day by flow cytometry. NAD(H)‐utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell‐free, ultrahigh‐throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per sample) carrying both variant DNA and an immobilised analogue of NAD+ were compartmentalised in water‐in‐oil emulsion droplets, together with cell‐free expression mixture and enzyme substrate, resulting in the recording of the phenotype on each bead. The beads’ phenotype could be read out and sorted for on a flow cytometer by using a highly sensitive fluorescent protein‐based sensor of the NAD+:NADH ratio. Integration of this “NAD‐display” approach with our previously described Split & Mix (SpliMLiB) method for generating large site‐saturation libraries allowed straightforward screening of fully balanced site saturation libraries of formate dehydrogenase, with diversities of 2×104. Based on modular design principles of synthetic biology NAD‐display offers access to sophisticated in vitro selections, avoiding complex technology platforms. A detection system was devised to screen dehydrogenase enzymes (DH) at ultrahigh‐throughput in in vitro droplet compartments. An NAD(H) analogue and a DH DNA library were co‐immobilised on beads. Water‐in‐oil emulsion droplets containing cell‐free expression mix and substrate allow compartmentalised protein production and catalysis. A fluorescent sensor of NAD(H) redox state was loaded onto beads, allowing bulk sorting of 2×104 DH variants in a day by flow cytometry. NAD(H)-utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell-free, ultrahigh-throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per sample) carrying both variant DNA and an immobilised analogue of NAD were compartmentalised in water-in-oil emulsion droplets, together with cell-free expression mixture and enzyme substrate, resulting in the recording of the phenotype on each bead. The beads' phenotype could be read out and sorted for on a flow cytometer by using a highly sensitive fluorescent protein-based sensor of the NAD :NADH ratio. Integration of this "NAD-display" approach with our previously described Split & Mix (SpliMLiB) method for generating large site-saturation libraries allowed straightforward screening of fully balanced site saturation libraries of formate dehydrogenase, with diversities of 2×10 . Based on modular design principles of synthetic biology NAD-display offers access to sophisticated in vitro selections, avoiding complex technology platforms. NAD(H)‐utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space, we demonstrate the utility of a cell‐free, ultrahigh‐throughput directed evolution platform for dehydrogenases. Microbeads (1.5 million per sample) carrying both variant DNA and an immobilised analogue of NAD+ were compartmentalised in water‐in‐oil emulsion droplets, together with cell‐free expression mixture and enzyme substrate, resulting in the recording of the phenotype on each bead. The beads’ phenotype could be read out and sorted for on a flow cytometer by using a highly sensitive fluorescent protein‐based sensor of the NAD+:NADH ratio. Integration of this “NAD‐display” approach with our previously described Split & Mix (SpliMLiB) method for generating large site‐saturation libraries allowed straightforward screening of fully balanced site saturation libraries of formate dehydrogenase, with diversities of 2×104. Based on modular design principles of synthetic biology NAD‐display offers access to sophisticated in vitro selections, avoiding complex technology platforms. |
| Author | Lindenburg, Laurens Hollfelder, Florian |
| AuthorAffiliation | 1 Department of Biochemistry University of Cambridge Tennis Court Road Cambridge CB2 1GA UK 2 Current address: Genmab Uppsalalaan 15 3584 CT Utrecht The Netherlands |
| AuthorAffiliation_xml | – name: 2 Current address: Genmab Uppsalalaan 15 3584 CT Utrecht The Netherlands – name: 1 Department of Biochemistry University of Cambridge Tennis Court Road Cambridge CB2 1GA UK |
| Author_xml | – sequence: 1 givenname: Laurens surname: Lindenburg fullname: Lindenburg, Laurens organization: Current address: Genmab – sequence: 2 givenname: Florian orcidid: 0000-0002-1367-6312 surname: Hollfelder fullname: Hollfelder, Florian email: fh111@cam.ac.uk organization: University of Cambridge |
| BackLink | https://www.ncbi.nlm.nih.gov/pubmed/33470025$$D View this record in MEDLINE/PubMed |
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| Cites_doi | 10.1016/j.tibs.2018.01.003 10.1016/j.chembiol.2012.06.009 10.1021/acssynbio.6b00188 10.1006/jmbi.1994.1244 10.1111/j.1432-1033.1980.tb04329.x 10.1111/j.1742-4658.2008.06533.x 10.1021/acs.analchem.8b01759 10.1021/ja00543a038 10.1021/acs.chemrev.8b00333 10.1016/j.jbiotec.2013.10.034 10.1007/978-1-4939-1053-3_7 10.1093/nar/gkaa270 10.1073/pnas.1700269114 10.1038/s41467-019-08441-5 10.1021/sb400110j 10.1016/j.ymben.2012.07.002 10.1002/anie.201001607 10.1021/acssynbio.8b00179 10.1073/pnas.1606927113 10.1073/pnas.0904764106 10.1016/j.cmet.2011.09.004 10.1073/pnas.102577799 10.1016/j.cmet.2011.08.012 10.1016/j.bbagen.2013.11.018 10.1007/s00253-012-4142-9 10.1038/nchembio.1385 10.1016/j.cbpa.2009.12.003 10.1002/cbic.201600484 10.1007/s10529-013-1156-z 10.1038/s41377-020-00358-9 10.1002/cbic.201402501 10.1038/nbt.4201 10.1021/acssynbio.9b00274 10.1039/C7CC05377K 10.1016/j.copbio.2016.04.023 10.1016/j.cmet.2015.04.009 10.1038/ncomms10008 10.1021/ac802613w 10.1016/j.sbi.2015.06.001 10.1016/j.cbpa.2017.02.018 10.1016/j.chembiol.2010.02.011 10.1002/ange.201001607 10.1016/j.sbi.2005.07.006 10.1042/bj3540455 10.1073/pnas.1115485109 10.1080/21655979.2014.1004020 10.1038/nmeth.4306 |
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| Keywords | cell-free expression formate dehydrogenase water-in-oil emulsion droplets saturation library directed evolution |
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| References | 2019; 8 2017; 6 2015; 6 1994; 237 2010; 14 2009; 81 2010; 17 2019; 10 2015; 33 2002; 99 2012; 19 2010 2010; 49 122 2011; 14 2012; 14 2016; 17 2018; 43 2014; 1840 2017; 114 2012; 109 2018; 7 2017; 53 2001; 354 2014; 3 2017; 37 2013; 10 2017; 14 2013; 35 2018; 118 2013; 97 2015; 21 2020; 9 2016; 113 2016; 42 2014; 15 2020; 48 2018; 90 2005; 15 2016; 29 2008; 275 2014; 1179 2014; 169 1980; 102 2018; 36 1980; 103 2009; 106 e_1_2_6_10_1 e_1_2_6_31_1 e_1_2_6_30_1 e_1_2_6_19_1 e_1_2_6_13_1 e_1_2_6_36_1 e_1_2_6_14_1 e_1_2_6_35_1 e_1_2_6_11_1 e_1_2_6_34_1 e_1_2_6_12_1 Cahn J. K. B. (e_1_2_6_32_1) 2016; 29 e_1_2_6_33_1 e_1_2_6_17_1 e_1_2_6_18_1 e_1_2_6_39_1 e_1_2_6_15_1 e_1_2_6_37_2 e_1_2_6_38_1 e_1_2_6_16_1 e_1_2_6_37_1 e_1_2_6_42_1 e_1_2_6_43_1 e_1_2_6_21_1 e_1_2_6_20_1 e_1_2_6_41_1 e_1_2_6_40_1 e_1_2_6_9_1 e_1_2_6_8_1 e_1_2_6_5_1 e_1_2_6_4_1 e_1_2_6_7_1 e_1_2_6_6_1 e_1_2_6_1_1 e_1_2_6_25_1 e_1_2_6_24_1 e_1_2_6_3_1 e_1_2_6_23_1 e_1_2_6_2_1 e_1_2_6_22_1 e_1_2_6_29_1 e_1_2_6_44_1 e_1_2_6_28_1 e_1_2_6_45_1 e_1_2_6_27_1 e_1_2_6_46_1 e_1_2_6_26_1 e_1_2_6_47_1 |
| References_xml | – volume: 113 start-page: E7383 year: 2016 end-page: E7389 publication-title: Proc. Natl. Acad. Sci. USA – volume: 102 start-page: 7104 year: 1980 end-page: 7105 publication-title: J. Am. Chem. Soc. – volume: 114 start-page: E2624 year: 2017 end-page: E2633 publication-title: Proc. Natl. Acad. Sci. USA – volume: 36 start-page: 880 year: 2018 end-page: 898 publication-title: Nat. Biotechnol. – volume: 3 start-page: 41 year: 2014 end-page: 47 publication-title: ACS Synth. Biol. – volume: 14 start-page: 122 year: 2010 end-page: 129 publication-title: Curr. Opin. Chem. Biol. – volume: 10 start-page: 711 year: 2019 publication-title: Nat. Commun. – volume: 169 start-page: 1 year: 2014 end-page: 8 publication-title: J. Biotechnol. – volume: 10 start-page: 50 year: 2013 end-page: 55 publication-title: Nat. Chem. Biol. – volume: 275 start-page: 3859 year: 2008 end-page: 3869 publication-title: FEBS J. – volume: 106 start-page: 15651 year: 2009 end-page: 15656 publication-title: Proc. Natl. Acad. Sci. USA – volume: 21 start-page: 777 year: 2015 end-page: 789 publication-title: Cell Metab. – volume: 118 start-page: 11707 year: 2018 end-page: 11794 publication-title: Chem. Rev. – volume: 37 start-page: 137 year: 2017 end-page: 144 publication-title: Curr. Opin. Chem. Biol. – volume: 97 start-page: 2473 year: 2013 end-page: 2481 publication-title: Appl. Microbiol. Biotechnol. – volume: 9 start-page: 118 year: 2020 publication-title: Light Sci. Appl. – volume: 99 start-page: 6597 year: 2002 end-page: 6602 publication-title: Proc. Natl. Acad. Sci. USA – volume: 15 start-page: 2710 year: 2014 end-page: 2718 publication-title: ChemBioChem – volume: 53 start-page: 9454 year: 2017 end-page: 9457 publication-title: Chem. Commun. – volume: 354 start-page: 455 year: 2001 end-page: 463 publication-title: Biochem. J. – volume: 237 start-page: 415 year: 1994 end-page: 422 publication-title: J. Mol. Biol. – volume: 42 start-page: 152 year: 2016 end-page: 158 publication-title: Curr. Opin. Biotechnol. – volume: 29 start-page: 31 year: 2016 end-page: 38 publication-title: Protein Eng. Des. Sel. – volume: 19 start-page: 1001 year: 2012 end-page: 1009 publication-title: Chem. Biol. – volume: 6 start-page: 106 year: 2015 end-page: 110 publication-title: Bioengineered – volume: 49 122 start-page: 5921 6057 year: 2010 2010 end-page: 5924 6060 publication-title: Angew. Chem. Int. Ed. Angew. Chem. – volume: 103 start-page: 421 year: 1980 end-page: 430 publication-title: Eur. J. Biochem. – volume: 7 start-page: 1715 year: 2018 end-page: 1721 publication-title: ACS Synth. Biol. – volume: 90 start-page: 9813 year: 2018 end-page: 9820 publication-title: Anal. Chem. – volume: 48 year: 2020 publication-title: Nucleic Acids Res. – volume: 14 start-page: 555 year: 2011 end-page: 566 publication-title: Cell Metab. – volume: 14 start-page: 720 year: 2017 end-page: 728 publication-title: Nat. Methods – volume: 17 start-page: 229 year: 2010 end-page: 235 publication-title: Chem. Biol. – volume: 1179 start-page: 103 year: 2014 end-page: 128 publication-title: Methods Mol. Biol. – volume: 33 start-page: 42 year: 2015 end-page: 51 publication-title: Curr. Opin. Struct. Biol. – volume: 109 start-page: E690 year: 2012 end-page: E697 publication-title: Proc. Natl. Acad. Sci. USA – volume: 43 start-page: 180 year: 2018 end-page: 198 publication-title: Trends Biochem. Sci. – volume: 35 start-page: 915 year: 2013 end-page: 919 publication-title: Biotechnol. Lett. – volume: 81 start-page: 5972 year: 2009 end-page: 5979 publication-title: Anal. Chem. – volume: 17 start-page: 2312 year: 2016 end-page: 2315 publication-title: ChemBioChem – volume: 1840 start-page: 951 year: 2014 end-page: 957 publication-title: Biochim. Biophys. Acta Gen. Subj. – volume: 8 start-page: 2690 year: 2019 end-page: 2700 publication-title: ACS Synth. Biol. – volume: 6 start-page: 326 year: 2017 end-page: 333 publication-title: ACS Synth. Biol. – volume: 15 start-page: 472 year: 2005 end-page: 478 publication-title: Curr. Opin. Struct. Biol. – volume: 14 start-page: 545 year: 2011 end-page: 554 publication-title: Cell Metab. – volume: 6 start-page: 10008 year: 2015 publication-title: Nat. Commun. – volume: 14 start-page: 504 year: 2012 end-page: 511 publication-title: Metab. Eng. – ident: e_1_2_6_1_1 doi: 10.1016/j.tibs.2018.01.003 – ident: e_1_2_6_4_1 doi: 10.1016/j.chembiol.2012.06.009 – ident: e_1_2_6_10_1 doi: 10.1021/acssynbio.6b00188 – ident: e_1_2_6_35_1 doi: 10.1006/jmbi.1994.1244 – ident: e_1_2_6_25_1 doi: 10.1111/j.1432-1033.1980.tb04329.x – ident: e_1_2_6_30_1 doi: 10.1111/j.1742-4658.2008.06533.x – ident: e_1_2_6_42_1 doi: 10.1021/acs.analchem.8b01759 – ident: e_1_2_6_24_1 doi: 10.1021/ja00543a038 – ident: e_1_2_6_6_1 doi: 10.1021/acs.chemrev.8b00333 – ident: e_1_2_6_18_1 doi: 10.1016/j.jbiotec.2013.10.034 – ident: e_1_2_6_43_1 doi: 10.1007/978-1-4939-1053-3_7 – ident: e_1_2_6_16_1 doi: 10.1093/nar/gkaa270 – ident: e_1_2_6_44_1 doi: 10.1073/pnas.1700269114 – ident: e_1_2_6_47_1 doi: 10.1038/s41467-019-08441-5 – ident: e_1_2_6_41_1 doi: 10.1021/sb400110j – ident: e_1_2_6_39_1 doi: 10.1016/j.ymben.2012.07.002 – ident: e_1_2_6_37_1 doi: 10.1002/anie.201001607 – ident: e_1_2_6_40_1 doi: 10.1021/acssynbio.8b00179 – ident: e_1_2_6_11_1 doi: 10.1073/pnas.1606927113 – ident: e_1_2_6_46_1 doi: 10.1073/pnas.0904764106 – ident: e_1_2_6_19_1 doi: 10.1016/j.cmet.2011.09.004 – ident: e_1_2_6_36_1 doi: 10.1073/pnas.102577799 – ident: e_1_2_6_20_1 doi: 10.1016/j.cmet.2011.08.012 – ident: e_1_2_6_21_1 doi: 10.1016/j.bbagen.2013.11.018 – ident: e_1_2_6_31_1 doi: 10.1007/s00253-012-4142-9 – ident: e_1_2_6_7_1 doi: 10.1038/nchembio.1385 – ident: e_1_2_6_8_1 doi: 10.1016/j.cbpa.2009.12.003 – ident: e_1_2_6_13_1 doi: 10.1002/cbic.201600484 – ident: e_1_2_6_26_1 doi: 10.1007/s10529-013-1156-z – ident: e_1_2_6_28_1 doi: 10.1038/s41377-020-00358-9 – ident: e_1_2_6_29_1 doi: 10.1002/cbic.201402501 – ident: e_1_2_6_33_1 doi: 10.1038/nbt.4201 – ident: e_1_2_6_34_1 doi: 10.1021/acssynbio.9b00274 – ident: e_1_2_6_12_1 doi: 10.1039/C7CC05377K – ident: e_1_2_6_9_1 doi: 10.1016/j.copbio.2016.04.023 – ident: e_1_2_6_14_1 doi: 10.1016/j.cmet.2015.04.009 – volume: 29 start-page: 31 year: 2016 ident: e_1_2_6_32_1 publication-title: Protein Eng. Des. Sel. – ident: e_1_2_6_5_1 doi: 10.1038/ncomms10008 – ident: e_1_2_6_22_1 doi: 10.1021/ac802613w – ident: e_1_2_6_2_1 doi: 10.1016/j.sbi.2015.06.001 – ident: e_1_2_6_3_1 doi: 10.1016/j.cbpa.2017.02.018 – ident: e_1_2_6_38_1 doi: 10.1016/j.chembiol.2010.02.011 – ident: e_1_2_6_37_2 doi: 10.1002/ange.201001607 – ident: e_1_2_6_15_1 doi: 10.1016/j.sbi.2005.07.006 – ident: e_1_2_6_23_1 doi: 10.1042/bj3540455 – ident: e_1_2_6_17_1 doi: 10.1073/pnas.1115485109 – ident: e_1_2_6_27_1 doi: 10.1080/21655979.2014.1004020 – ident: e_1_2_6_45_1 doi: 10.1038/nmeth.4306 |
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| Snippet | NAD(H)‐utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space,... NAD(H)-utiliing enzymes have been the subject of directed evolution campaigns to improve their function. To enable access to a larger swath of sequence space,... |
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| SubjectTerms | Beads Biological evolution cell-free expression Dehydrogenases Directed evolution Flow cytometry Fluorescence Formate dehydrogenase Libraries Microspheres Modular design NAD NADH Nanoparticles Nicotinamide adenine dinucleotide Phenotypes Saturation saturation library Screening Substrates water-in-oil emulsion droplets |
| Title | “NAD‐display”: Ultrahigh‐Throughput in Vitro Screening of NAD(H) Dehydrogenases Using Bead Display and Flow Cytometry |
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