Functional Expression of the Aromatic Prenyltransferase NphB in Chlamydomonas reinhardtii Highlights Challenges in Cannabinoid Biocatalysis

Nouemssi, Serge Basile; Bouhadada, Ayoub; Beauchemin, Rémy; Custeau, Alexandre; Gélinas, Sarah-Ève; Merindol, Natacha; Meddeb-Mouelhi, Fatma; Germain, Hugo; Desgagné-Penix, Isabel

doi:10.3390/catal16040346

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Functional Expression of the Aromatic Prenyltransferase NphB in Chlamydomonas reinhardtii Highlights Challenges in Cannabinoid Biocatalysis

by

Serge Basile Nouemssi

,

Ayoub Bouhadada

,

Rémy Beauchemin

,

Alexandre Custeau

,

Sarah-Ève Gélinas

,

Natacha Merindol

,

Fatma Meddeb-Mouelhi

,

Hugo Germain

and

Isabel Desgagné-Penix

^*

Department of Biochemistry, Chemistry, Physics, and Forensic Science, Université du Québec à Trois-Rivières, 3351 Boulevard des Forges, Trois-Rivières, QC G9A 5H7, Canada

^*

Author to whom correspondence should be addressed.

Catalysts 2026, 16(4), 346; https://doi.org/10.3390/catal16040346

Submission received: 13 March 2026 / Revised: 2 April 2026 / Accepted: 10 April 2026 / Published: 13 April 2026

(This article belongs to the Special Issue Biocatalysis and Biosynthesis: Opportunities and Challenges)

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Abstract

Cannabinoids are high-value bioactive compounds whose sustainable production remains challenging, prompting interest in biocatalytic and microbial platforms as alternatives to plant extraction. In this study, we investigated the heterologous expression and functionality of two key cannabinoid-related enzymes in the photosynthetic microalga Chlamydomonas reinhardtii: the aromatic prenyltransferase, NphB_G286S/Y288A from Streptomyces sp., and the plant-derived cannabidiolic acid synthase (CBDAS) from Cannabis sativa. Codon-optimized genes were introduced into the nuclear genome of C. reinhardtii using several construct configurations and promoters, and stable transformants were generated and characterized for genomic integration, transcript accumulation, protein production, enzymatic activity, and cannabinoid-related metabolite formation. While NphB protein accumulation was achieved under the PSAD promoter control, CBDAS was not detected at the protein level under any condition tested. In vitro enzymatic assays using soluble algal protein extracts from NphB-expressing lines confirmed catalytic activity, yielding cannabigerolic acid (CBGA), reaching up to 633 ± 58 µg L⁻¹. However, no CBGA production was detected in vivo, despite substrate supplementation. These results indicate that, although bacterial prenyltransferase can be functionally expressed in C. reinhardtii, efficient metabolic conversion in vivo is limited by cellular and biochemical constraints, including substrate availability, intracellular compartmentalization, and potential competition with endogenous pathways. In contrast, the absence of detectable CBDAS highlights the challenges associated with expressing complex plant oxidocyclases in this photosynthetic host. Overall, this work provides mechanistic insights into enzyme compatibility and metabolic bottlenecks in microalgal systems and outlines key considerations for the future development of photosynthetic platforms for cannabinoid biocatalysis.

Keywords: microalgae biofactories; cannabinoids; photosynthetic biocatalyst; metabolic engineering; aromatic prenyltransferase; synthetic biology; cannabidiolic acid synthase

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MDPI and ACS Style

Nouemssi, S.B.; Bouhadada, A.; Beauchemin, R.; Custeau, A.; Gélinas, S.-È.; Merindol, N.; Meddeb-Mouelhi, F.; Germain, H.; Desgagné-Penix, I. Functional Expression of the Aromatic Prenyltransferase NphB in Chlamydomonas reinhardtii Highlights Challenges in Cannabinoid Biocatalysis. Catalysts 2026, 16, 346. https://doi.org/10.3390/catal16040346

AMA Style

Nouemssi SB, Bouhadada A, Beauchemin R, Custeau A, Gélinas S-È, Merindol N, Meddeb-Mouelhi F, Germain H, Desgagné-Penix I. Functional Expression of the Aromatic Prenyltransferase NphB in Chlamydomonas reinhardtii Highlights Challenges in Cannabinoid Biocatalysis. Catalysts. 2026; 16(4):346. https://doi.org/10.3390/catal16040346

Chicago/Turabian Style

Nouemssi, Serge Basile, Ayoub Bouhadada, Rémy Beauchemin, Alexandre Custeau, Sarah-Ève Gélinas, Natacha Merindol, Fatma Meddeb-Mouelhi, Hugo Germain, and Isabel Desgagné-Penix. 2026. "Functional Expression of the Aromatic Prenyltransferase NphB in Chlamydomonas reinhardtii Highlights Challenges in Cannabinoid Biocatalysis" Catalysts 16, no. 4: 346. https://doi.org/10.3390/catal16040346

APA Style

Nouemssi, S. B., Bouhadada, A., Beauchemin, R., Custeau, A., Gélinas, S.-È., Merindol, N., Meddeb-Mouelhi, F., Germain, H., & Desgagné-Penix, I. (2026). Functional Expression of the Aromatic Prenyltransferase NphB in Chlamydomonas reinhardtii Highlights Challenges in Cannabinoid Biocatalysis. Catalysts, 16(4), 346. https://doi.org/10.3390/catal16040346

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Functional Expression of the Aromatic Prenyltransferase NphB in Chlamydomonas reinhardtii Highlights Challenges in Cannabinoid Biocatalysis

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