{
  "abstract": "Anisole is a valuable aromatic ether and promising green solvent whose industrial manufacture relies exclusively on petroleum-derived phenol and hazardous alkali-mediated methylation. Here, we establish the first de novo microbial biosynthesis of anisole from glucose in Pseudomonas putida KT2440 by integrating retrosynthetic pathway design with growth-coupled directed evolution. Using a selection platform in an engineered double auxotroph, in which growth depends on S-adenosylmethionine-dependent methylation, we evolved a promiscuous plant chavicol O-methyltransferase to yield phenol O-methyltransferase 1 (POMT1; bearing a key D264N substitution), which improved anisole production by 2.3-fold over the wild-type enzyme. To overcome product volatility, we integrated an in situ two-phase extraction system using dodecane that achieved near-quantitative product recovery. Then, we compared two phenol routes from the shikimate pathway, selected the 4-hydroxybenzoate route, and raised precursor supply by deleting competing reactions and introducing feedback-resistant and reduced-flux variants in two shikimate pathway enzymes. The final strain produced up to 0.57 mM anisole (62 mg/L) from glucose.",
  "abstract_shown": true,
  "affiliations": [
    "Wageningen University & Research"
  ],
  "article_type": "New results",
  "arxiv": null,
  "author_total": 6,
  "authors": [
    {
      "affs": [
        0
      ],
      "name": "L. Bruinsma"
    },
    {
      "name": "D. Rosas Gomez"
    },
    {
      "name": "C. La Rocca"
    },
    {
      "name": "J. E. Escrig Molina"
    },
    {
      "name": "V. Martins dos Santos"
    },
    {
      "name": "C. Batianis"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/37n7rc6j1mdj5.bib",
  "cited_by": 0,
  "doi": "10.64898/2026.11.02.763652",
  "fulltext_url": "https://www.biorxiv.org/content/10.64898/2026.11.02.763652v1.full",
  "id": "37n7rc6j1mdj5",
  "issns": [],
  "landing_url": "https://www.biorxiv.org/content/10.64898/2026.11.02.763652v1",
  "language": null,
  "licence": "CC-BY-NC-ND",
  "licence_label": "CC BY-NC-ND",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by-nc-nd/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "biorxiv",
      "bioengineering"
    ]
  ],
  "openalex": null,
  "pdf_url": "https://www.biorxiv.org/content/10.64898/2026.11.02.763652v1.full.pdf",
  "pmcid": null,
  "pmid": null,
  "published": "2026-11-02",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/37n7rc6j1mdj5.ris",
  "sources": [
    {
      "first_seen": "2026-11-04T00:21:33Z",
      "native_id": "10.64898/2026.11.02.763652",
      "source": "biorxiv",
      "url": "https://www.biorxiv.org/content/10.64898/2026.11.02.763652v1"
    }
  ],
  "subjects": [
    {
      "field": "biology",
      "origin": "biorxiv:bioengineering",
      "subfield": "systems-synthetic"
    }
  ],
  "title": "Establishing de novo microbial biosynthesis of anisole",
  "title_text": "Establishing de novo microbial biosynthesis of anisole",
  "type": "preprint",
  "updated_at": "2026-11-05T00:20:56Z",
  "url": "https://research.draconitic.com/paper/37n7rc6j1mdj5",
  "venue": "bioRxiv",
  "versions": []
}