{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,23]],"date-time":"2026-09-23T04:01:20Z","timestamp":1790136080786,"version":"4.0.1"},"reference-count":78,"publisher":"Wiley","issue":"9","license":[{"start":{"date-parts":[[2023,4,7]],"date-time":"2023-04-07T00:00:00Z","timestamp":1680825600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"content-domain":{"domain":["chemistry-europe.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["ChemBioChem"],"published-print":{"date-parts":[[2023,5,2]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Whereas directed evolution and rational design by structural inspection are established tools for enzyme redesign, computational methods are less mature but have the potential to predict small sets of mutants with desired properties without laboratory screening of large libraries. We have explored the use of computational enzyme redesign to change the enantioselectivity of a highly thermostable alcohol dehydrogenase from <jats:italic>Thermus thermophilus<\/jats:italic> in the asymmetric reduction of ketones. The enzyme reduces acetophenone to (<jats:italic>S<\/jats:italic>)\u20101\u2010phenylethanol. To invert the enantioselectivity, we used an adapted CASCO workflow which included Rosetta for enzyme design and molecular dynamics simulations for ranking. To correct for unrealistic binding modes, we used Boltzmann weighing of binding energies computed by a linear interaction energy approach. This computationally cheap method predicted four variants with inverted enantioselectivity, each with 6\u20138 mutations around the substrate\u2010binding site, causing only modest reduction (2\u2010 to 7\u2010fold) of <jats:italic>k<jats:sub>cat<\/jats:sub><\/jats:italic>\/<jats:italic>K<\/jats:italic><jats:sub>M<\/jats:sub> values. Laboratory testing showed that three variants indeed had inverted enantioselectivity, producing (<jats:italic>R<\/jats:italic>)\u2010alcohols with up to 99\u2009% enantiomeric excess. The broad substrate range allowed reduction of acetophenone derivatives with full conversion to highly enantioenriched alcohols. The results demonstrate the use of computational methods to control ketoreductase stereoselectivity in asymmetric transformations with minimal experimental screening.<\/jats:p>","DOI":"10.1002\/cbic.202300032","type":"journal-article","created":{"date-parts":[[2023,3,14]],"date-time":"2023-03-14T04:19:45Z","timestamp":1678767585000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":13,"title":["Computationally Supported Inversion of Ketoreductase Stereoselectivity"],"prefix":"10.1002","volume":"24","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2115-4047","authenticated-orcid":false,"given":"Estela","family":"Delgado\u2010Arciniega","sequence":"first","affiliation":[{"name":"Biotransformation and Biocatalysis Groningen Biomolecular Sciences and Biotechnology Institute (GBB) University of Groningen  Nijenborgh 4 9747 AG Groningen The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0891-6972","authenticated-orcid":false,"given":"Hein J.","family":"Wijma","sequence":"additional","affiliation":[{"name":"Biotransformation and Biocatalysis Groningen Biomolecular Sciences and Biotechnology Institute (GBB) University of Groningen  Nijenborgh 4 9747 AG Groningen The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chantal","family":"Hummel","sequence":"additional","affiliation":[{"name":"Biotransformation and Biocatalysis Groningen Biomolecular Sciences and Biotechnology Institute (GBB) University of Groningen  Nijenborgh 4 9747 AG Groningen The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0834-2043","authenticated-orcid":false,"given":"Dick B.","family":"Janssen","sequence":"additional","affiliation":[{"name":"Biotransformation and Biocatalysis Groningen Biomolecular Sciences and Biotechnology Institute (GBB) University of Groningen  Nijenborgh 4 9747 AG Groningen The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2023,4,7]]},"reference":[{"key":"e_1_2_11_1_1","doi-asserted-by":"publisher","DOI":"10.1002\/anie.202006648"},{"key":"e_1_2_11_1_2","doi-asserted-by":"publisher","DOI":"10.1002\/ange.202006648"},{"key":"e_1_2_11_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.bmc.2017.05.023"},{"key":"e_1_2_11_3_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.cbpa.2009.12.003"},{"key":"e_1_2_11_4_1","doi-asserted-by":"publisher","DOI":"10.1002\/anie.200502746"},{"key":"e_1_2_11_4_2","doi-asserted-by":"publisher","DOI":"10.1002\/ange.200502746"},{"key":"e_1_2_11_5_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.tet.2005.10.044"},{"key":"e_1_2_11_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.enzmictec.2019.03.001"},{"key":"e_1_2_11_7_1","doi-asserted-by":"publisher","DOI":"10.1021\/acscatal.8b03382"},{"key":"e_1_2_11_8_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.cbpa.2013.01.017"},{"key":"e_1_2_11_9_1","doi-asserted-by":"publisher","DOI":"10.1021\/acscatal.5b02752"},{"key":"e_1_2_11_10_1","doi-asserted-by":"publisher","DOI":"10.1021\/cs501457v"},{"key":"e_1_2_11_11_1","doi-asserted-by":"publisher","DOI":"10.1021\/ja974129t"},{"key":"e_1_2_11_12_1","doi-asserted-by":"publisher","DOI":"10.1007\/3-540-68116-7_2"},{"key":"e_1_2_11_13_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.biortech.2017.03.096"},{"key":"e_1_2_11_14_1","doi-asserted-by":"publisher","DOI":"10.1021\/ie503060a"},{"key":"e_1_2_11_15_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.tetasy.2005.02.030"},{"key":"e_1_2_11_16_1","doi-asserted-by":"publisher","DOI":"10.1006\/jmbi.2000.3758"},{"key":"e_1_2_11_17_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.251555398"},{"key":"e_1_2_11_18_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcatb.2011.01.017"},{"key":"e_1_2_11_19_1","doi-asserted-by":"publisher","DOI":"10.1002\/pro.462"},{"key":"e_1_2_11_20_1","doi-asserted-by":"publisher","DOI":"10.1021\/ci500126x"},{"key":"e_1_2_11_21_1","doi-asserted-by":"publisher","DOI":"10.1039\/C4OB00794H"},{"key":"e_1_2_11_22_1","doi-asserted-by":"publisher","DOI":"10.1021\/ol702638j"},{"key":"e_1_2_11_23_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbiotec.2015.11.003"},{"key":"e_1_2_11_24_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-016-0028-x"},{"key":"e_1_2_11_25_1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0019230"},{"key":"e_1_2_11_26_1","doi-asserted-by":"publisher","DOI":"10.1021\/cb3006227"},{"key":"e_1_2_11_27_1","doi-asserted-by":"publisher","DOI":"10.1002\/pro.5560070314"},{"key":"e_1_2_11_28_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1507910112"},{"key":"e_1_2_11_29_1","doi-asserted-by":"publisher","DOI":"10.1021\/cr050283j"},{"key":"e_1_2_11_30_1","doi-asserted-by":"publisher","DOI":"10.1002\/anie.201411415"},{"key":"e_1_2_11_30_2","doi-asserted-by":"publisher","DOI":"10.1002\/ange.201411415"},{"key":"e_1_2_11_31_1","doi-asserted-by":"publisher","DOI":"10.1002\/jcc.21546"},{"key":"e_1_2_11_32_1","doi-asserted-by":"publisher","DOI":"10.1002\/cbic.201900726"},{"key":"e_1_2_11_33_1","doi-asserted-by":"publisher","DOI":"10.1046\/j.1432-1033.2002.03130.x"},{"key":"e_1_2_11_34_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0167-7799(98)01207-4"},{"key":"e_1_2_11_35_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-020-10440-2"},{"key":"e_1_2_11_36_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi051596b"},{"key":"e_1_2_11_37_1","doi-asserted-by":"publisher","DOI":"10.1021\/op9002714"},{"key":"e_1_2_11_38_1","doi-asserted-by":"publisher","DOI":"10.1007\/BFb0103304"},{"key":"e_1_2_11_39_1","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.00217-08"},{"key":"e_1_2_11_40_1","doi-asserted-by":"publisher","DOI":"10.2174\/092986610790963564"},{"key":"e_1_2_11_41_1","doi-asserted-by":"publisher","DOI":"10.1093\/protein\/7.3.385"},{"key":"e_1_2_11_42_1","doi-asserted-by":"publisher","DOI":"10.1021\/jm7012198"},{"key":"e_1_2_11_43_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00894-015-2883-y"},{"key":"e_1_2_11_44_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.bpj.2010.02.034"},{"key":"e_1_2_11_45_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.cbi.2008.09.018"},{"key":"e_1_2_11_46_1","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btu426"},{"key":"e_1_2_11_47_1","doi-asserted-by":"publisher","DOI":"10.1021\/j100142a004"},{"key":"e_1_2_11_48_1","unstructured":"D.\u2005J.\u2005F.\u2005M.\u2005J. Frisch G.\u2005W. Trucks H.\u2005B. Schlegel G.\u2005E. Scuseria M.\u2005A. Robb J.\u2005R. Cheeseman G. Scalmani V. Barone G.\u2005A. Petersson H. Nakatsuji X. Li M. Caricato A. Marenich J. Bloino B.\u2005G. Janesko R. Gomperts B. Mennucci H.\u2005P. Hratchian J.\u2005V. Ort Gaussian Inc. Wallingford CT2009."},{"key":"e_1_2_11_49_1","doi-asserted-by":"publisher","DOI":"10.1021\/jm0608210"},{"key":"e_1_2_11_50_1","unstructured":"D.\u2005A. Case R.\u2005M. Betz D.\u2005S. Cerutti T.\u2005E. Cheatham\u2005III T.\u2005A. Darden R.\u2005E. Duke T.\u2005J. Giese H. Gohlke A.\u2005W. Goetz N. Homeyer S. Izadi P. Janowski J. Kaus A. Kovalenko T.\u2005S. Lee S. LeGrand P. Li C. Lin T. Luchko R. Luo B. Madej D. Mermelstein K.\u2005M. Merz G. Monard H. Nguyen H.\u2005T. Nguyen I. Omelyan A. Onufriev D.\u2005R. Roe A. Roitberg C. Sagui C.\u2005L. Simmerling W.\u2005M. Botello-Smith J. Swails R.\u2005C. Walker J. Wang R.\u2005M. Wolf X. Wu L. Xiao P.\u2005A. Kollman AMBER 2016 University of California San Francisco 2016."},{"key":"e_1_2_11_51_1","doi-asserted-by":"publisher","DOI":"10.1002\/jcc.23899"},{"key":"e_1_2_11_52_1","doi-asserted-by":"publisher","DOI":"10.1002\/prot.20251"},{"key":"e_1_2_11_53_1","doi-asserted-by":"publisher","DOI":"10.1063\/1.470117"},{"key":"e_1_2_11_54_1","doi-asserted-by":"publisher","DOI":"10.1023\/A:1007930623000"},{"key":"e_1_2_11_55_1","doi-asserted-by":"publisher","DOI":"10.1002\/(SICI)1097-0134(19990215)34:3<395::AID-PROT11>3.0.CO;2-4"},{"key":"e_1_2_11_56_1","doi-asserted-by":"publisher","DOI":"10.1021\/acs.jcim.7b00222"},{"key":"e_1_2_11_57_1","unstructured":"R Core Team \u201cR: A Language and Environment for Statistical Computing \u201d can be found underhttps:\/\/www.r-project.org 2022."},{"key":"e_1_2_11_58_1","unstructured":"Stan Development Team \u201cRStan: the R interface to Stan.\u201d R package version 2.21.7 \u201d can be found under https:\/\/mc-stan.org\/.2022."},{"key":"e_1_2_11_59_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.21168"},{"key":"e_1_2_11_60_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ab.2006.07.027"},{"key":"e_1_2_11_61_1","doi-asserted-by":"publisher","DOI":"10.1021\/acscatal.9b05223"},{"key":"e_1_2_11_62_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0022-2836(03)00081-0"},{"key":"e_1_2_11_63_1","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.71.10.5728-5734.2005"},{"key":"e_1_2_11_64_1","doi-asserted-by":"publisher","DOI":"10.1021\/acscatal.8b00807"},{"key":"e_1_2_11_65_1","doi-asserted-by":"publisher","DOI":"10.1021\/jacs.8b08640"},{"key":"e_1_2_11_66_1","doi-asserted-by":"publisher","DOI":"10.1021\/ja00007a002"},{"key":"e_1_2_11_67_1","volume-title":"Structure and Mechanism in Protein Science. A Guide to Enzyme Catalysis and Protein Folding","author":"Fersht A.","year":"1999"},{"key":"e_1_2_11_68_1","doi-asserted-by":"publisher","DOI":"10.1515\/hsz-2018-0299"},{"key":"e_1_2_11_69_1","doi-asserted-by":"publisher","DOI":"10.1002\/anie.200353055"},{"key":"e_1_2_11_69_2","doi-asserted-by":"publisher","DOI":"10.1002\/ange.200353055"},{"key":"e_1_2_11_70_1","doi-asserted-by":"publisher","DOI":"10.1021\/acscatal.7b01933"},{"key":"e_1_2_11_71_1","doi-asserted-by":"publisher","DOI":"10.1002\/prot.24463"},{"key":"e_1_2_11_72_1","doi-asserted-by":"publisher","DOI":"10.1002\/anie.202205735"},{"key":"e_1_2_11_73_1","doi-asserted-by":"publisher","DOI":"10.1039\/C3OB42057D"},{"key":"e_1_2_11_74_1","doi-asserted-by":"publisher","DOI":"10.1039\/c2ob06870b"}],"container-title":["ChemBioChem"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/cbic.202300032","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/chemistry-europe.onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/cbic.202300032","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T13:46:27Z","timestamp":1760276787000},"score":1,"resource":{"primary":{"URL":"https:\/\/chemistry-europe.onlinelibrary.wiley.com\/doi\/10.1002\/cbic.202300032"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,4,7]]},"references-count":78,"journal-issue":{"issue":"9","published-print":{"date-parts":[[2023,5,2]]}},"alternative-id":["10.1002\/cbic.202300032"],"URL":"https:\/\/doi.org\/10.1002\/cbic.202300032","archive":["Portico"],"relation":{},"ISSN":["1439-4227","1439-7633"],"issn-type":[{"value":"1439-4227","type":"print"},{"value":"1439-7633","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,4,7]]},"assertion":[{"value":"2023-01-14","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2023-04-07","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e202300032"}}