Metabolic Engineering for Efficient Ketocarotenoid Accumulation in the Green Microalga Chlamydomonas reinhardtii

by Sofia Amendola, Jacob S. Kneip, Florian Meyer, Federico Perozeni, Stefano Cazzaniga, Kyle J. Lauersen, Matteo Ballottari, Thomas Baier
Article Year: 2023 DOI: 10.1021/acssynbio.2c00616

Bibliography

Amendola, S., Kneip, J.S., Meyer, F., Perozeni, F., Cazzaniga, S., Lauersen, K.J., Ballottari, M., Baier, T. (2023) Metabolic Engineering for Efficient Ketocarotenoid Accumulation in the Green Microalga Chlamydomonas reinhardtii. ACS Synthetic Biology.

Abstract

Astaxanthin is a valuable ketocarotenoid with various pharmaceutical and nutraceutical applications. Green microalgae harbor natural capacities for pigment accumulation due to their 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway. Recently, a redesigned ß-carotene ketolase (BKT) was found to enable ketocarotenoid accumulation in the model microalga Chlamydomonas reinhardtii, and transformants exhibited reduced photoinhibition under high-light. Here, a systematic screening by synthetic transgene design of carotenoid pathway enzymes and overexpression from the nuclear genome identified phytoene synthase (PSY/crtB) as a bottleneck for carotenoid accumulation in C. reinhardtii. Increased ß-carotene hydroxylase (CHYB) activity was found to be essential for engineered astaxanthin accumulation. A combined BKT, crtB, and CHYB expression strategy resulted in a volumetric astaxanthin production of 9.5 ± 0.3 mg L–1 (4.5 ± 0.1 mg g–1 CDW) in mixotrophic and 23.5 mg L–1 (1.09 mg L–1 h–1) in high cell density conditions, a 4-fold increase compared to previous reports in C. reinhardtii. This work presents a systematic investigation of bottlenecks in astaxanthin accumulation in C. reinhardtii and the phototrophic green cell factory design for competitive use in industrial biotechnology.

Keywords

Biosynthesis Dyes and pigments Genetics Hydrocarbons Peptides and proteins