{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T01:52:33Z","timestamp":1760061153825,"version":"build-2065373602"},"reference-count":35,"publisher":"Wiley","issue":"8","license":[{"start":{"date-parts":[[2020,6,3]],"date-time":"2020-06-03T00:00:00Z","timestamp":1591142400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["21476011"],"award-info":[{"award-number":["21476011"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["analyticalsciencejournals.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Biotech &amp; Bioengineering"],"published-print":{"date-parts":[[2020,8]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Most expression systems are tailored for model organisms rather than nonmodel organisms. However, heterologous gene expression in model organisms constrains the innate advantages of original strain carrying gene of interest. In this study, T7 expression system was developed in nonmodel bacterium <jats:italic>Klebsiella pneumoniae<\/jats:italic> for production of chemicals. First, we engineered a recombinant <jats:italic>K. pneumoniae<\/jats:italic> strain harboring two vectors. One vector was used to express T7 RNA polymerase (T7 RNAP) which would drive the expression of <jats:italic>egfp<\/jats:italic> in the other vector. This recombinant strain demonstrated 15.73\u2010fold of fluorescence relative to wild\u2010type <jats:italic>K. pneumoniae<\/jats:italic> and showed similar level of fluorescence to recombinant <jats:italic>Escherichia coli<\/jats:italic> overexpressing <jats:italic>egfp<\/jats:italic>. When <jats:italic>egfp<\/jats:italic> was replaced by <jats:italic>puuC<\/jats:italic>, an endogenous aldehyde dehydrogenase catalyzing 3\u2010hydroxypropionic acid (3\u2010HP) biosynthesis in <jats:italic>K. pneumoniae<\/jats:italic>, the recombinant strain coexpressing T7 RNAP and PuuC showed high\u2010level PuuC expression. In shake\u2010flask cultivation, this recombinant strain produced 1.72\u2009g\/L 3\u2010HP in 24\u2009hr, which was 3.24 times that of wild\u2010type <jats:italic>K. pneumoniae<\/jats:italic> (0.53\u2009g\/L). To mitigate plasmid burden, the vector expressing T7 RNAP was eliminated, but the T7 RNAP expression cassette was integrated into <jats:italic>K. pneumoniae<\/jats:italic> genome. The resulting strain harboring only PuuC expression vector produced 2.44\u2009g\/L 3\u2010HP in 24\u2009hr under shake\u2010flask conditions, which was 1.46 times that of the strain harboring both T7 RNAP and PuuC expression vectors. In bioreactor cultivation, this strain generated 67.59\u2009g\/L 3\u2010HP and did not show significantly halted growth. Overall, these results indicate that the engineered T7 expression system functioned efficiently in <jats:italic>K. pneumoniae<\/jats:italic>. This study provides a paradigm for the development of T7 expression system in prokaryotes.<\/jats:p>","DOI":"10.1002\/bit.27434","type":"journal-article","created":{"date-parts":[[2020,5,21]],"date-time":"2020-05-21T08:51:14Z","timestamp":1590051074000},"page":"2446-2459","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Development of orthogonal T7 expression system in <i>Klebsiella pneumoniae<\/i>"],"prefix":"10.1002","volume":"117","author":[{"given":"Peng","family":"Zhao","sequence":"first","affiliation":[{"name":"Beijing Key Laboratory of Bioprocess, College of Life Science and Technology Beijing University of Chemical Technology  Beijing China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Minrui","family":"Ren","sequence":"additional","affiliation":[{"name":"Beijing Key Laboratory of Bioprocess, College of Life Science and Technology Beijing University of Chemical Technology  Beijing China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xizhen","family":"Ge","sequence":"additional","affiliation":[{"name":"College of Biochemical Engineering Beijing Union University  Beijing China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5261-9516","authenticated-orcid":false,"given":"Pingfang","family":"Tian","sequence":"additional","affiliation":[{"name":"Beijing Key Laboratory of Bioprocess, College of Life Science and Technology Beijing University of Chemical Technology  Beijing China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tianwei","family":"Tan","sequence":"additional","affiliation":[{"name":"Beijing Key Laboratory of Bioprocess, College of Life Science and Technology Beijing University of Chemical Technology  Beijing 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