{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,21]],"date-time":"2026-08-21T13:15:06Z","timestamp":1787318106558,"version":"build-2736575974"},"reference-count":35,"publisher":"Wiley","issue":"7","license":[{"start":{"date-parts":[[2025,4,8]],"date-time":"2025-04-08T00:00:00Z","timestamp":1744070400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":["analyticalsciencejournals.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Biotech &amp; Bioengineering"],"published-print":{"date-parts":[[2025,7]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>\n                    Transaminase (TA)\u2010catalyzed asymmetric amination is considered as a green chemistry approach to synthesize pharmaceutical analogs, but their ability to accept substrate for catalyzing sterically hindered ketones remains a challenge. Sitagliptin is an antihyperglycemic drug to treat type II diabetes. Herein, we exploited an efficient (\n                    <jats:italic>R<\/jats:italic>\n                    )\u2010selective TA to biosynthesize sitagliptin analog (\n                    <jats:italic>R<\/jats:italic>\n                    )\u20103\u2010amino\u20101\u2010morpholino\u20104\u2010(2,4,5\u2010trifluorophenyl)butan\u20101\u2010one. Starting from a previously constructed (\n                    <jats:italic>R<\/jats:italic>\n                    )\u2010ATA5, two rounds of directed evolution were performed through combining error\u2010prone PCR, site\u2010directed saturation and combinatorial mutagenesis. The resultant variant ATA5\/F189H\/S236T\/M121H showed a 10.2\u2010fold higher activity and a 4\u2010fold improved half\u2010life at 45\u00b0C. Crucially, the variant was able to either catalyze the amination of 700\u2009mM substrate with a conversion up to 93.1% and product\n                    <jats:italic>e.e<\/jats:italic>\n                    .&gt;\u200999% in a cosolvent reaction system, or biotransform 200\u2009mM substrate with a conversion of 97.6% and product\n                    <jats:italic>e.e<\/jats:italic>\n                    .&gt;\u200999% in a cosolvent\u2010free system. Furthermore, the structural analysis gave insight into how the mutations affected enzymatic activity and thermostability. This study, which consists of constructing a robust (\n                    <jats:italic>R<\/jats:italic>\n                    )\u2010selective TA and the new synthesis route with the highest conversion ever reported, provides a reference for industrial manufacturing sitagliptin analog.\n                  <\/jats:p>","DOI":"10.1002\/bit.28988","type":"journal-article","created":{"date-parts":[[2025,4,8]],"date-time":"2025-04-08T09:47:54Z","timestamp":1744105674000},"page":"1735-1746","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["Directed Evolution of an (\n                    <i>R<\/i>\n                    )\u2010Selective Transaminase Toward Higher Efficiency of Sitagliptin Analog Biosynthesis"],"prefix":"10.1002","volume":"122","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-2433-0816","authenticated-orcid":false,"given":"Dong\u2010Xu","family":"Jia","sequence":"first","affiliation":[{"name":"Key Laboratory of Bioorganic Synthesis of Zhejiang Province College of Biotechnology and Bioengineering, Zhejiang University of Technology Hangzhou China"},{"name":"The National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals Zhejiang University of Technology Hangzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lei","family":"Zang","sequence":"additional","affiliation":[{"name":"Key Laboratory of Bioorganic Synthesis of Zhejiang Province College of Biotechnology and Bioengineering, Zhejiang University of Technology Hangzhou China"},{"name":"The National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals Zhejiang University of Technology Hangzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chi\u2010De","family":"Ni","sequence":"additional","affiliation":[{"name":"Key Laboratory of Bioorganic Synthesis of Zhejiang Province College of Biotechnology and Bioengineering, Zhejiang University of Technology Hangzhou China"},{"name":"The National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals Zhejiang University of Technology Hangzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jia\u2010Le","family":"Wang","sequence":"additional","affiliation":[{"name":"Key Laboratory of Bioorganic Synthesis of Zhejiang Province College of Biotechnology and Bioengineering, Zhejiang University of Technology Hangzhou China"},{"name":"The National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals Zhejiang University of Technology Hangzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hai","family":"Yu","sequence":"additional","affiliation":[{"name":"Key Laboratory of Bioorganic Synthesis of Zhejiang Province College of Biotechnology and Bioengineering, Zhejiang University of Technology Hangzhou China"},{"name":"The National and Local Joint Engineering Research 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