{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,14]],"date-time":"2026-02-14T07:03:04Z","timestamp":1771052584701,"version":"3.50.1"},"reference-count":72,"publisher":"Wiley","issue":"3","license":[{"start":{"date-parts":[[2025,12,2]],"date-time":"2025-12-02T00:00:00Z","timestamp":1764633600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"},{"start":{"date-parts":[[2025,12,2]],"date-time":"2025-12-02T00:00:00Z","timestamp":1764633600000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"content-domain":{"domain":["analyticalsciencejournals.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Biotech &amp; Bioengineering"],"published-print":{"date-parts":[[2026,3]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>\n                    Toluene\n                    <jats:italic>o<\/jats:italic>\n                    \u2010xylene monooxygenase (ToMO) of\n                    <jats:italic>Pseudomonas<\/jats:italic>\n                    sp. OX1 was investigated as a drug\u2010metabolizing enzyme for the first time and was found to metabolize chlorzoxazone and resveratrol to form human metabolites 6\u2010chlorzoxazone (0.045\u2009\u00b1\u20090.016\u2009nmol\/hr\/mg protein) and piceatannol (0.014\u2009\u00b1\u20090.009\u2009nmol\/hr\/mg protein), respectively, though at low rates. ToMO also forms 2\u2010acetamidophenol (2\u2010AAP, 27%), 3\u2010AAP (42%), and 4\u2010AAP (31%) from acetanilide at 3.6\u2009\u00b1\u20090.3\u2009nmol\/hr\/mg protein. Multiple\u2010site saturation mutagenesis at positions I100\/E103\/A107 of the alpha\u2010subunit along with site\u2010directed mutagenesis approaches were used to isolate thirty\u2010seven different ToMO variants with enhanced activities and\/or fine\u2010tuned specificities. Specifically, variant I100V\/E103T was identified with 2.1\u2010 and 49\u2010fold higher activities towards acetanilide and chlorzoxazone, respectively, compared to native ToMO. Variant I100V\/E103T also had the regiospecificity of acetanilide change from 31% to 100% 4\u2010AAP, mimicking human liver enzyme behavior. In addition, several variants showed up to 3.7\u2010, 1.6\u2010, and 3.2\u2010fold improved selectivity for 2\u2010, 3\u2010, and 4\u2010AAP formation, respectively. For resveratrol, variant I100T\/E103L was a better catalyst than native ToMO, exhibiting 34\u2010fold higher activity. The results presented here demonstrate the potential of nonhuman ToMO variants in drug metabolism and contribute to the list of research on probing this promising enzyme.\n                  <\/jats:p>","DOI":"10.1002\/bit.70117","type":"journal-article","created":{"date-parts":[[2025,12,2]],"date-time":"2025-12-02T10:57:31Z","timestamp":1764673051000},"page":"657-669","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Drug Metabolism by Engineered Toluene\n                    <i>o<\/i>\n                    \u2010Xylene Monooxygenases of\n                    <i>Pseudomonas<\/i>\n                    sp. OX1"],"prefix":"10.1002","volume":"123","author":[{"given":"Noella","family":"Younan","sequence":"first","affiliation":[{"name":"Department of Chemistry California State University Stanislaus Turlock California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Felice A.","family":"Dacpano","sequence":"additional","affiliation":[{"name":"Department of Chemistry California State University Stanislaus Turlock California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Eli","family":"Frazer","sequence":"additional","affiliation":[{"name":"Department of Chemistry California State University Stanislaus Turlock California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Angeline","family":"Dauz","sequence":"additional","affiliation":[{"name":"Department of Chemistry California State University Stanislaus Turlock California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Areli","family":"Tlatelpa","sequence":"additional","affiliation":[{"name":"Department of Chemistry California State University Stanislaus Turlock California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0858-3170","authenticated-orcid":false,"given":"G\u00f6n\u00fcl","family":"Vardar\u2010Schara","sequence":"additional","affiliation":[{"name":"Department of Chemistry California State University Stanislaus Turlock California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2025,12,2]]},"reference":[{"key":"e_1_2_7_2_1","doi-asserted-by":"publisher","DOI":"10.1002\/bies.202100052"},{"key":"e_1_2_7_3_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi901150a"},{"key":"e_1_2_7_4_1","doi-asserted-by":"publisher","DOI":"10.1021\/acs.chemrestox.6b00396"},{"key":"e_1_2_7_5_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-014-5767-7"},{"key":"e_1_2_7_6_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1527-3458.2006.00250.x"},{"key":"e_1_2_7_7_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0022-3565(25)03462-7"},{"key":"e_1_2_7_8_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M113.479717"},{"key":"e_1_2_7_9_1","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.71.8.4744-4750.2005"},{"key":"e_1_2_7_10_1","doi-asserted-by":"publisher","DOI":"10.1128\/JB.184.2.344-349.2002"},{"key":"e_1_2_7_11_1","unstructured":"Corso G. 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