Biocatalytic Production of Galantamine in Yeast through Cytochrome P450 Optimization

Abstract

Galantamine is a pharmaceutically relevant Amaryllidaceae alkaloid used for the treatment of Alzheimer's disease. Its structural complexity and low abundance in plants motivate the development of alternative manufacturing routes. Here, we established the first engineered yeast platform for the biocatalytic production of galantamine from 4OMe-norbelladine. Heterologous expression of the downstream pathway enzymes NtCYP96T6, NtNMT1, and NtAKR1 from Narcissus cv. Tete-a-Tete in Saccharomyces cerevisiae was complemented by optimization of cultivation temperature, carbon source, and medium pH enabling the first demonstration of galantamine production in yeast. Systematic screening of cytochrome P450 reductase and cytochrome b partners to boost NtCYP96T6 activity led to the identification of a novel reductase mined from the Narcissus pseudonarcissus transcriptome, NpCPR, which supported the highest pathway flux and yielded 7.0 {+/-} 0.6 mg/L galantamine, corresponding to a 7.6 % molar yield from 250 M 4OMe-norbelladine and an approximately 173-fold improvement over the parental strain. We further exploited this yeast cell factory for the precursor-directed biosynthesis of 7F-galantamine, highlighting the potential of pathway enzyme promiscuity to access new-to-nature GAL analogues that may be challenging to produce through conventional chemical synthesis. Together, this work establishes a foundation for microbial galantamine production and biosynthetic diversification of its pharmaceutically relevant scaffold.

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BibTeX RIS

BibTeX

@misc{holtz2026biocatalytic,
  title = {{Biocatalytic Production of Galantamine in Yeast through Cytochrome P450 Optimization}},
  author = {M. Holtz and S. de Haan and C. Castellvi Domingo and N. G. Madsen and M. D. F. Salazar and F. G. Hansson and A. C. A. van Aalst and J. A. Arnesen and C. Crocoll and S. d'Oelsnitz and M. K. Jensen and C. G. Acevedo-Rocha},
  howpublished = {bioRxiv},
  year = {2026},
  month = sep,
  doi = {10.64898/2026.09.09.750345},
  url = {https://www.biorxiv.org/content/10.64898/2026.09.09.750345v1},
  note = {CC BY-NC-ND},
}

RIS

TY  - UNPB
TI  - Biocatalytic Production of Galantamine in Yeast through Cytochrome P450 Optimization
AU  - M. Holtz
AU  - S. de Haan
AU  - C. Castellvi Domingo
AU  - N. G. Madsen
AU  - M. D. F. Salazar
AU  - F. G. Hansson
AU  - A. C. A. van Aalst
AU  - J. A. Arnesen
AU  - C. Crocoll
AU  - S. d'Oelsnitz
AU  - M. K. Jensen
AU  - C. G. Acevedo-Rocha
PY  - 2026
DA  - 2026/09/09
PB  - bioRxiv
DO  - 10.64898/2026.09.09.750345
UR  - https://www.biorxiv.org/content/10.64898/2026.09.09.750345v1
AB  - Galantamine is a pharmaceutically relevant Amaryllidaceae alkaloid used for the treatment of Alzheimer's disease. Its structural complexity and low abundance in plants motivate the development of alternative manufacturing routes. Here, we established the first engineered yeast platform for the biocatalytic production of galantamine from 4OMe-norbelladine. Heterologous expression of the downstream pathway enzymes NtCYP96T6, NtNMT1, and NtAKR1 from Narcissus cv. Tete-a-Tete in Saccharomyces cerevisiae was complemented by optimization of cultivation temperature, carbon source, and medium pH enabling the first demonstration of galantamine production in yeast. Systematic screening of cytochrome P450 reductase and cytochrome b partners to boost NtCYP96T6 activity led to the identification of a novel reductase mined from the Narcissus pseudonarcissus transcriptome, NpCPR, which supported the highest pathway flux and yielded 7.0 {+/-} 0.6 mg/L galantamine, corresponding to a 7.6 % molar yield from 250 M 4OMe-norbelladine and an approximately 173-fold improvement over the parental strain. We further exploited this yeast cell factory for the precursor-directed biosynthesis of 7F-galantamine, highlighting the potential of pathway enzyme promiscuity to access new-to-nature GAL analogues that may be challenging to produce through conventional chemical synthesis. Together, this work establishes a foundation for microbial galantamine production and biosynthetic diversification of its pharmaceutically relevant scaffold.
ER  -

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